Proliferative Capacity of Single Isolated CD34+ Hematopoietic Stem/Progenitor Cells in Paroxysmal Nocturnal Hemoglobinuria

Paroxysmal nocturnal hemoglobinuria (PNH) results from somatic mutations of the X-linked PIG-A (phosphatidylinositol glycan-class A) gene, which occurs on a hematopoietic stem cell level, leading to a proportion of blood cells being deficient in all glycosylphosphatidylinositol (GPI)-anchored surface proteins. Although these GPI-deficient cells can explain many of the clinical symptoms of PNH, the pathogenesis of PNH is still somewhat obscure and many questions remain. To assess the hematopoietic defect involved in PNH, CD34+ CD59+ (normal phenotype hematopoietic stem/progenitor) and CD34+ CD59- (PNH phenotype) cells from PNH patients (n = 16) and CD34+ CD59+ cells from healthy volunteers (n = 10) were sorted as single cells into 96-well flat-bottom culture plates containing culture medium supplemented with stem cell factor, interleukin (IL)-3, erythropoietin, granulocyte-macrophage-colony-stimulating factor (GM-CSF), G-CSF, IL-6, thrombopoietin, and Flt-3 ligand. We found that the single PNH CD34+ CD59- cells had a growth advantage over the single CD34+ CD59+ cells to some extent, but they both had impaired growth abilities compared with CD34+ cells from healthy volunteers.     

Increased CD4+CD8+ Double Positive T Cells during Hantaan Virus Infection

Hantaan virus (HTNV) infection causes an epidemic of hemorrhagic fever with renal syndrome (HFRS) mainly in Asia. It is well known that T cells mediated anti-viral immune response. Although previous studies showed that double positive T (DP T) cells, a little portion of T lymphocytes, were involved in adaptive immune response during virus infection, their kinetic changes and roles in HTNV infection have not yet been explored. In this study, we characterized DP T cells from HFRS patients based on flow cytometry data combined with scRNA-seq data. We showed that HTNV infection caused the upregulation of DP T cells in the peripheral blood, which were correlated with disease stage. The scRNA-seq data clustered DP T cells, unraveled their gene expression profile, and estimated the ordering of these cells. The production of granzyme B and CD107a from DP T cells and the abundant TCR distribution indicated the anti-viral property of DP T cells. In conclusion, this study identified, for the first time, an accumulation of DP T cells in the peripheral blood of HFRS patients and suggested these DP T cells belonging to CD8⁺T cells lineage. The DP T cells shared the similar characteristics with cytotoxic T cells (CTL) and exerted an anti-viral role in HFRS.     

Ethanol Suppression of the Functional State of Polymorphonuclear Leukocytes Obtained From Uninfected and Simian Immunodeficiency Virus Infected Rhesus Macaques

BACKGROUND: Human immunodeficiency virus (HIV) infection and alcohol abuse frequently coexist in the host and are known to suppress individually the host response to a variety of opportunistic infections. METHODS: This study examined the effects of in vitro ethanol exposure on several functions of polymorphonuclear leukocytes (PMNs) that were obtained from uninfected and simian immunodeficiency virus (SIV)-infected rhesus macaques, at the asymptomatic and terminal stages of infection. RESULTS: The PMNs obtained from rhesus macaques at both the asymptomatic and terminal stage of SIV disease had elevated phagocytic activity and increased CD11b expression compared with PMNs from uninfected animals. In vitro 100 mM ethanol suppressed phagocytosis and CD11b adhesion molecule expression by PMNs, regardless of the stage of SIV infection. Treatment of PMNs with granulocyte colony-stimulating factor (G-CSF) attenuated the inhibitory effect seen with prior ethanol exposure. CONCLUSIONS: These data demonstrate that the functional state of PMNs from uninfected as well as SIV-infected rhesus macaques is impaired by direct exposure to intoxicating concentrations of ethanol and that this effect can be attenuated by G-CSF. If alcohol intoxication similarly suppressed PMN function in vivo, it would further increase susceptibility of these hosts to secondary infections. Furthermore, G-CSF may be useful in overcoming the suppressive effects of ethanol on PMN function in such patients.     

Two New Pseudopod Morphologies Displayed by the Human Hematopoietic KG1a Progenitor Cell Line and by Primary Human CD34+ Cells

A primitive human hematopoietic myeloid progenitor cell line, KG1a,characterized by high expression of the CD34 surface antigen has beenobserved to extend long, thin pseudopodia. Once extended, thesepseudopods may take on one of two newly described morphologies, tenupodia or magnupodia. Tenupodia are very thin and form in linear segments. They adhere to the substrate, can bifurcate multiple times,and often appear to connect the membranes of cells more than 300 µmapart. Magnupodia are much thicker and have been observed to extendmore than 330 µm away from the cell. Magnupods are flexible and canexhibit rapid dynamic motion, extending or retracting in a few seconds.During retraction, the extended material often pools into a bulblocated on the pod. Both morphologies can adhere to substrates coatedwith fibronectin, collagen IV, and laminin as well as plastic. The CD34and CD44 antigens are also present on the surface of these podia.Primary human CD34⁺ cells from fetal liver, umbilicalcord blood, adult bone marrow, and mobilized peripheral blood extendthese podia as well. The morphology that these pseudopods exhibitsuggest that they may play both sensory and mechanical roles duringcell migration and homing after bone marrow transplantation.     

Expansion of CD4+CD25+FoxP3+ Regulatory T Cells in Chronic Hepatitis C Virus Infection

Background: Regulatory T cells (Tregs) have been involved in impaired immunity and may have a pivotal role in persistence of viral infections. Objective: To develop a simple and reliable in-house three color flow cytometery of peripheral blood to understand the role of HCV infection in the increase of Tregs. Methods: The level of naturally occurring CD4+CD25+FoxP3+ regulatory T cells (nTregs) in 20 chronically infected with hepatitis C virus (HCV) patients was compared to those of 15 healthy individuals by flowcytometry. In a different approach we performed permeabilization and intracellular staining before surface staining which allows the preservation of the surface molecules in the combined detection process and results in the normal frequency of nTregs in blood. Results: Using the optimized method, it was shown that a significantly higher proportion of nTregs in the total CD4+ T cell population was seen in the peripheral blood of chronic HCV patients (0.83 ± 0.21%, p=0.05) as compared to controls (0.26 ± 0.1, p=0.05). Conclusions: In accordance with other studies, we showed that HCV infection induces a dramatic increase in Tregs, which might contribute to the immune response failure during HCV infection.     

Inverse Correlation Between Circulating Endothelial Progenitor Cells With CD34+CD133+ and the Severity of Coronary Atherosclerosis Assessed by Syntax Score

BackgroundThe number of endothelial progenitor cells (EPCs) descends when atherosclerosis developed. The objective was to compare the number of CD34 + CD133 + cells with the severity of atherosclerosis assessed by Syntax score.MethodsThe study included 80 patients with stable angina undergoing coronary angiography. Patients were classified into single-vessel group, multiple-vessel group and normal group according to angiography. The percentage of CD34 + CD133 + cells in the mononuclear cells isolated from peripheral blood of different groups by flow cytometric analysis was compared. The quantity of CD34 + CD133 + EPCs was log transformed to improve normality (lgEPC). Syntax score was used in this study to assess the extent of coronary artery disease.Results:The level of lgEPC was lower in the single-vessel group than that in the normal group (− 3.42 ± 0.44 versus − 3.17 ± 0.39, P < 0.05), and the level of lgEPC was lower in the multiple-vessel group than that in the single vessel group (− 3.63 ± 0.31 versus − 3.42 ± 0.44, P < 0.05). An inverse correlation between lgEPC and Syntax score analyzed by linear regression.ConclusionsEPC level probably serves as a predictor of the development and severity of atherosclerosis on a cellular level. EPC, a relatively more important risk factor, perhaps protects against coronary artery disease.     

Hepatitis D Virus-Specific CD8+ T Cells Have a Memory-Like Phenotype Associated With Viral Immune Escape in Patients With Chronic Hepatitis D Virus Infection

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The CD34 Molecule and Hematopoietic Progenitor Cell Studies - a Challenge in Clinical Medicine

The CD34 molecule, which is expressed at various levels on virtually all human hematopoietic progenitor cells (HPC), is a heavy glycosylated molecule with many different epitopes. A large series of CD34-specific monoclonal antibodies specific for different epitopes has been generated. Some of these antibodies do not react with particular HPC subpopulations. Thus, the antibodies used for HPC enumeration and isolation have to be carefully chosen with respect to what is aimed at.