Engineered living blood vessels: functional endothelia generated from human umbilical cord-derived progenitors.

PubMed

Schmidt, Dörthe; Asmis, Lars M; Odermatt, Bernhard; Kelm, Jens; Breymann, Christian; Gössi, Matthias; Genoni, Michele; Zund, Gregor; Hoerstrup, Simon P

2006-10-01

Tissue-engineered living blood vessels (TEBV) with growth capacity represent a promising new option for the repair of congenital malformations. We investigate the functionality of TEBV with endothelia generated from human umbilical cord blood-derived endothelial progenitor cells. Tissue-engineered living blood vessels were generated from human umbilical cord-derived myofibroblasts seeded on biodegradable vascular scaffolds, followed by endothelialization with differentiated cord blood-derived endothelial progenitor cells. During in vitro maturation the TEBV were exposed to physiologic conditioning in a flow bioreactor. For functional assessment, a subgroup of TEBV was stimulated with tumor necrosis factor-alpha. Control vessels endothelialized with standard vascular endothelial cells were treated in parallel. Analysis of the TEBV included histology, immunohistochemistry, biochemistry (extracellular matrix analysis, DNA), and biomechanical testing. Endothelia were analyzed by flow cytometry and immunohistochemistry (CD31, von Willebrand factor, thrombomodulin, tissue factor, endothelial nitric oxide synthase). Histologically, a three-layered tissue organization of the TEBV analogous to native vessels was observed, and biochemistry revealed the major matrix constituents (collagen, proteoglycans) of blood vessels. Biomechanical properties (Young's modulus, 2.03 +/- 0.65 MPa) showed profiles resembling those of native tissue. Endothelial progenitor cells expressed typical endothelial cell markers CD31, von Willebrand factor, and endothelial nitric oxide synthase comparable to standard vascular endothelial cells. Stimulation with tumor necrosis factor-alpha resulted in physiologic upregulation of tissue factor and downregulation of thrombomodulin expression. These results indicate that TEBV with tissue architecture and functional endothelia similar to native blood vessels can be successfully generated from human umbilical cord progenitor cells. Thus, blood-derived progenitor cells obtained before or at birth may enable the clinical realization of tissue engineering constructs for pediatric applications.

Red blood cells in sports: effects of exercise and training on oxygen supply by red blood cells

PubMed Central

Mairbäurl, Heimo

2013-01-01

During exercise the cardiovascular system has to warrant substrate supply to working muscle. The main function of red blood cells in exercise is the transport of O2 from the lungs to the tissues and the delivery of metabolically produced CO2 to the lungs for expiration. Hemoglobin also contributes to the blood's buffering capacity, and ATP and NO release from red blood cells contributes to vasodilation and improved blood flow to working muscle. These functions require adequate amounts of red blood cells in circulation. Trained athletes, particularly in endurance sports, have a decreased hematocrit, which is sometimes called “sports anemia.” This is not anemia in a clinical sense, because athletes have in fact an increased total mass of red blood cells and hemoglobin in circulation relative to sedentary individuals. The slight decrease in hematocrit by training is brought about by an increased plasma volume (PV). The mechanisms that increase total red blood cell mass by training are not understood fully. Despite stimulated erythropoiesis, exercise can decrease the red blood cell mass by intravascular hemolysis mainly of senescent red blood cells, which is caused by mechanical rupture when red blood cells pass through capillaries in contracting muscles, and by compression of red cells e.g., in foot soles during running or in hand palms in weightlifters. Together, these adjustments cause a decrease in the average age of the population of circulating red blood cells in trained athletes. These younger red cells are characterized by improved oxygen release and deformability, both of which also improve tissue oxygen supply during exercise. PMID:24273518

Cellular function reinstitution of offspring red blood cells cloned from the sickle cell disease patient blood post CRISPR genome editing.

PubMed

Wen, Jianguo; Tao, Wenjing; Hao, Suyang; Zu, Youli

2017-06-13

Sickle cell disease (SCD) is a disorder of red blood cells (RBCs) expressing abnormal hemoglobin-S (HbS) due to genetic inheritance of homologous HbS gene. However, people with the sickle cell trait (SCT) carry a single allele of HbS and do not usually suffer from SCD symptoms, thus providing a rationale to treat SCD. To validate gene therapy potential, hematopoietic stem cells were isolated from the SCD patient blood and treated with CRISPR/Cas9 approach. To precisely dissect genome-editing effects, erythroid progenitor cells were cloned from single colonies of CRISPR-treated cells and then expanded for simultaneous gene, protein, and cellular function studies. Genotyping and sequencing analysis revealed that the genome-edited erythroid progenitor colonies were converted to SCT genotype from SCD genotype. HPLC protein assays confirmed reinstallation of normal hemoglobin at a similar level with HbS in the cloned genome-edited erythroid progenitor cells. For cell function evaluation, in vitro RBC differentiation of the cloned erythroid progenitor cells was induced. As expected, cell sickling assays indicated function reinstitution of the genome-edited offspring SCD RBCs, which became more resistant to sickling under hypoxia condition. This study is an exploration of genome editing of SCD HSPCs.

Hematopoietic Colony Formation from Human Growth Factor-Dependent TF1 Cells and Human Cord Blood Myeloid Progenitor Cells Depends on SHP2 Phosphatase Function

PubMed Central

Etienne-Julan, Maryse; Gotoh, Akihiko; Braun, Stephen E.; Lu, Li; Cooper, Scott; Feng, Gen-Sheng; Li, Xing Jun

2013-01-01

The protein tyrosine phosphatase, SHP2, is widely expressed; however, previous studies demonstrated that hematopoietic cell development more stringently requires Shp2 expression compared to other tissues. Furthermore, somatic gain-of-function SHP2 mutants are commonly found in human myeloid leukemias. Given that pharmacologic inhibitors to SHP2 phosphatase activity are currently in development as putative antileukemic agents, we conducted a series of experiments examining the necessity of SHP2 phosphatase activity for human hematopoiesis. Anti-sense oligonucleotides to human SHP2 coding sequences reduced human cord blood- and human cell line, TF1-derived colony formation. Expression of truncated SHP2 bearing its Src homology 2 (SH2) domains, but lacking the phosphatase domain similarly reduced human cord blood- and TF1-derived colony formation. Mechanistically, expression of truncated SHP2 reduced the interaction between endogenous, full-length SHP2 with the adapter protein, Grb2. To verify the role of SHP2 phosphatase function in human hematopoietic cell development, human cord blood CD34+ cells were transduced with a leukemia-associated phosphatase gain-of-function SHP2 mutant or with a phosphatase dead SHP2 mutant, which indicated that increased phosphatase function enhanced, while decreased SHP2 phosphatase function reduced, human cord blood-derived colonies. Collectively, these findings indicate that SHP2 phosphatase function regulates human hematopoietic cell development and imply that the phosphatase component of SHP2 may serve as a pharmacologic target in human leukemias bearing increased SHP2 phosphatase activity. PMID:23082805

Hematopoietic colony formation from human growth factor-dependent TF1 cells and human cord blood myeloid progenitor cells depends on SHP2 phosphatase function.

PubMed

Broxmeyer, Hal E; Etienne-Julan, Maryse; Gotoh, Akihiko; Braun, Stephen E; Lu, Li; Cooper, Scott; Feng, Gen-Sheng; Li, Xing Jun; Chan, Rebecca J

2013-03-15

The protein tyrosine phosphatase, SHP2, is widely expressed; however, previous studies demonstrated that hematopoietic cell development more stringently requires Shp2 expression compared to other tissues. Furthermore, somatic gain-of-function SHP2 mutants are commonly found in human myeloid leukemias. Given that pharmacologic inhibitors to SHP2 phosphatase activity are currently in development as putative antileukemic agents, we conducted a series of experiments examining the necessity of SHP2 phosphatase activity for human hematopoiesis. Anti-sense oligonucleotides to human SHP2 coding sequences reduced human cord blood- and human cell line, TF1-derived colony formation. Expression of truncated SHP2 bearing its Src homology 2 (SH2) domains, but lacking the phosphatase domain similarly reduced human cord blood- and TF1-derived colony formation. Mechanistically, expression of truncated SHP2 reduced the interaction between endogenous, full-length SHP2 with the adapter protein, Grb2. To verify the role of SHP2 phosphatase function in human hematopoietic cell development, human cord blood CD34+ cells were transduced with a leukemia-associated phosphatase gain-of-function SHP2 mutant or with a phosphatase dead SHP2 mutant, which indicated that increased phosphatase function enhanced, while decreased SHP2 phosphatase function reduced, human cord blood-derived colonies. Collectively, these findings indicate that SHP2 phosphatase function regulates human hematopoietic cell development and imply that the phosphatase component of SHP2 may serve as a pharmacologic target in human leukemias bearing increased SHP2 phosphatase activity.

Engineering Robust and Functional Vascular Networks in Vivo with Human Adult and Cord Blood-Derived Progenitor Cells

DTIC Science & Technology

2008-12-01

for other sources of ECs such as those derived from embryonic and adult progenitor cells ( Rafii ; Lyden 2003). For example, human ES-derived...functional endothelial precursors. Blood, 95, 952-958. Rafii , S., and D. Lyden, 2003: Therapeutic stem and progenitor cell transplantation for

Synthesis and characterisation of glucose-functional glycopolymers and gold nanoparticles: study of their potential interactions with ovine red blood cells.

PubMed

Wilkins, Laura E; Phillips, Daniel J; Deller, Robert C; Davies, Gemma-Louise; Gibson, Matthew I

2015-03-20

Carbohydrate-protein interactions can assist with the targeting of polymer- and nano-delivery systems. However, some potential protein targets are not specific to a single cell type, resulting in reductions in their efficacy due to undesirable non-specific cellular interactions. The glucose transporter 1 (GLUT-1) is expressed to different extents on most cells in the vasculature, including human red blood cells and on cancerous tissue. Glycosylated nanomaterials bearing glucose (or related) carbohydrates, therefore, could potentially undergo unwanted interactions with these transporters, which may compromise the nanomaterial function or lead to cell agglutination, for example. Here, RAFT polymerisation is employed to obtain well-defined glucose-functional glycopolymers as well as glycosylated gold nanoparticles. Agglutination and binding assays did not reveal any significant binding to ovine red blood cells, nor any haemolysis. These data suggest that gluco-functional nanomaterials are compatible with blood, and their lack of undesirable interactions highlights their potential for delivery and imaging applications. Copyright © 2014 Elsevier Ltd. All rights reserved.

A novel perivascular cell population in the zebrafish brain.

PubMed

Venero Galanternik, Marina; Castranova, Daniel; Gore, Aniket V; Blewett, Nathan H; Jung, Hyun Min; Stratman, Amber N; Kirby, Martha R; Iben, James; Miller, Mayumi F; Kawakami, Koichi; Maraia, Richard J; Weinstein, Brant M

2017-04-11

The blood-brain barrier is essential for the proper homeostasis and function of the CNS, but its mechanism of function is poorly understood. Perivascular cells surrounding brain blood vessels are thought to be important for blood-brain barrier establishment, but their roles are not well defined. Here, we describe a novel perivascular cell population closely associated with blood vessels on the zebrafish brain. Based on similarities in their morphology, location, and scavenger behavior, these cells appear to be the zebrafish equivalent of cells variably characterized as Fluorescent Granular Perithelial cells (FGPs), perivascular macrophages, or 'Mato Cells' in mammals. Despite their macrophage-like morphology and perivascular location, zebrafish FGPs appear molecularly most similar to lymphatic endothelium, and our imaging studies suggest that these cells emerge by differentiation from endothelium of the optic choroidal vascular plexus. Our findings provide the first report of a perivascular cell population in the brain derived from vascular endothelium.

Cord blood derived CD4+ CD25(high) T cells become functional regulatory T cells upon antigen encounter.

PubMed

Mayer, Elisabeth; Bannert, Christina; Gruber, Saskia; Klunker, Sven; Spittler, Andreas; Akdis, Cezmi A; Szépfalusi, Zsolt; Eiwegger, Thomas

2012-01-01

Upon antigen exposure, cord blood derived T cells respond to ubiquitous environmental antigens by high proliferation. To date it remains unclear whether these "excessive" responses relate to different regulatory properties of the putative T regulatory cell (Treg) compartment or even expansion of the Treg compartment itself. Cord blood (>37 week of gestation) and peripheral blood (healthy controls) were obtained and different Treg cell subsets were isolated. The suppressive potential of Treg populations after antigen exposure was evaluated via functional inhibition assays ([(3)H]thymidine incorporation assay and CFSE staining) with or without allergen stimulation. The frequency and markers of CD4(+)CD25(high)FoxP3(+) T cells were characterized by mRNA analysis and flow cytometry. Cord blood derived CD4(+)CD25(high) cells did not show substantial suppressor capacity upon TCR activation, in contrast to CD4(+)CD25(high) cells freshly purified from adult blood. This could not be explained by a lower frequency of FoxP3(+)CD4(+)CD25(high)cells or FOXP3 mRNA expression. However, after antigen-specific stimulation in vitro, these cells showed strong proliferation and expansion and gained potent suppressive properties. The efficiency of their suppressive capacity can be enhanced in the presence of endotoxins. If T-cells were sorted according to their CD127 expression, a tiny subset of Treg cells (CD4(+)CD25(+)CD127(low)) is highly suppressive even without prior antigen exposure. Cord blood harbors a very small subset of CD4(+)CD25(high) Treg cells that requires antigen-stimulation to show expansion and become functional suppressive Tregs.

Novel methods of treating ovarian infertility in older and POF women, testicular infertility, and other human functional diseases.

PubMed

Bukovsky, Antonin

2015-02-25

In vitro maturation (IVM) and in vitro fertilization (IVF) technologies are facing with growing demands of older women to conceive. Although ovarian stem cells (OSCs) of older women are capable of producing in vitro fresh oocyte-like cells (OLCs), such cells cannot respond to IVM and IVF due to the lack of granulosa cells required for their maturation. Follicular renewal is also dependent on support of circulating blood mononuclear cells. They induce intermediary stages of meiosis (metaphase I chromosomal duplication and crossover, anaphase, telophase, and cytokinesis) in newly emerging ovarian germ cells, as for the first time demonstrated here, induce formation of granulosa cells, and stimulate follicular growth and development. A pretreatment of OSC culture with mononuclear cells collected from blood of a young healthy fertile woman may cause differentiation of bipotential OSCs into both developing germ and granulosa cells. A small blood volume replacement may enable treatment of ovarian infertility in vivo. The transferred mononuclear cells may temporarily rejuvenate virtually all tissues, including improvement of the function of endocrine tissues. Formation of new follicles and their development may be sufficient for IVM and IVF. The novel proposed in vitro approaches may be used as a second possibility. Infertility of human males affects almost a half of the infertility cases worldwide. Small blood volume replacement from young healthy fertile men may also be easy approach for the improvement of sperm quality in older or other affected men. In addition, body rejuvenation by small blood volume replacement from young healthy individuals of the same sex could represent a decline of in vitro methodology in favor of in vivo treatment for human functional diseases. Here we propose for the first time that blood mononuclear cells are essential for rejuvenation of those tissues, where immune system components participate in an appropriate division and differentiation of tissue stem cells. If needed, small blood volume replacement from distinct young healthy individuals could be utilized in six month intervals for repair of young altered or aged reproductive and other tissue functions. Systemic and local use of honey bee propolis tincture is an alternative option for functional rejuvenation of some tissues.

Involvement of blood mononuclear cells in the infertility, age-associated diseases and cancer treatment

PubMed Central

Bukovsky, Antonin

2016-01-01

Blood mononuclear cells consist of T cells and monocyte derived cells. Beside immunity, the blood mononuclear cells belong to the complex tissue control system (TCS), where they exhibit morphostatic function by stimulating proliferation of tissue stem cells followed by cellular differentiation, that is stopped after attaining the proper functional stage, which differs among various tissue types. Therefore, the term immune and morphostatic system (IMS) should be implied. The TCS-mediated morphostasis also consists of vascular pericytes controlled by autonomic innervation, which is regulating the quantity of distinct tissues in vivo. Lack of proper differentiation of tissue cells by TCS causes either tissue underdevelopment, e.g., muscular dystrophy, or degenerative functional failures, e.g., type 1 diabetes and age-associated diseases. With the gradual IMS regression after 35 years of age the gonadal infertility develops, followed by a growing incidence of age-associated diseases and cancers. Without restoring an altered TCS function in a degenerative disease, the implantation of tissue-specific stem cells alone by regenerative medicine can not be successful. Transfused young blood could temporarily restore fertility to enable parenthood. The young blood could also temporarily alleviate aging diseases, and this can be extended by substances inducing IMS regeneration, like the honey bee propolis. The local and/or systemic use of honey bee propolis stopped hair and teeth loss, regressed varicose veins, improved altered hearing, and lowered high blood pressure and sugar levels. Complete regression of stage IV ovarian cancer with liver metastases after a simple elaborated immunotherapy is also reported. PMID:28074124

Phenotypic, ultra-structural and functional characterization of bovine peripheral blood dendritic cell subsets

USDA-ARS?s Scientific Manuscript database

Dendritic cells (DC) are multifunctional cells that bridge the gap between innate and adaptive immune systems. In bovine, significant information is lacking on the precise identity and role of peripheral blood DC subsets. In this study, we identify and characterize bovine peripheral blood DC subsets...

An affordable method to obtain cultured endothelial cells from peripheral blood

PubMed Central

Bueno-Betí, Carlos; Novella, Susana; Lázaro-Franco, Macarena; Pérez-Cremades, Daniel; Heras, Magda; Sanchís, Juan; Hermenegildo, Carlos

2013-01-01

The culture of endothelial progenitor cells (EPC) provides an excellent tool to research on EPC biology and vascular regeneration and vasculogenesis. The use of different protocols to obtain EPC cultures makes it difficult to obtain comparable results in different groups. This work offers a systematic comparison of the main variables of most commonly used protocols for EPC isolation, culture and functional evaluation. Peripheral blood samples from healthy individuals were recovered and mononuclear cells were cultured. Different recovery and culture conditions were tested: blood volume, blood anticoagulant, coating matrix and percentage of foetal bovine serum (FBS) in culture media. The success of culture procedure, first colonies of endothelial cells appearance time, correlation with number of circulating EPC (cEPC) and functional comparison with human umbilical vein endothelial cells (HUVEC) were studied. The use of heparin, a minimum blood volume of 30 ml, fibronectin as a coating matrix and endothelial growing media-2 supplemented with 20% FBS increased the success of obtaining EPC cultures up to 80% of the processed samples while reducing EPC colony appearance mean time to a minimum of 13 days. Blood samples exhibiting higher cEPC numbers resulted in reduced EPC colony appearance mean time. Cells isolated by using this combination were endothelial cell-like EPCs morphological and phenotypically. Functionally, cultured EPC showed decreased growing and vasculogenic capacity when compared to HUVEC. Thus, above-mentioned conditions allow the isolation and culture of EPC with smaller blood volumes and shorter times than currently used protocols. PMID:24118735

Altered phenotype and function of NK cells infiltrating human papillomavirus (HPV)-associated genital warts during HIV infection.

PubMed

Bere, Alfred; Tayib, Shahila; Kriek, Jean-Mari; Masson, Lindi; Jaumdally, Shameem Z; Barnabas, Shaun L; Carr, William H; Allan, Bruce; Williamson, Anna-Lise; Denny, Lynette; Passmore, Jo-Ann S

2014-02-01

HIV-infected individuals experience more persistent HPV infections and are less likely to resolve genital warts. This study compared phenotype and functions of NK and T cells from genital warts and blood from 67 women. We compared in vitro functional responses of NK and T cells by multiparametric flow cytometry. HIV+ women had significantly lower frequencies of CD4 T cells in warts (p = 0.001) and blood (p = 0.001). While the distribution of NK cell subsets was similar, HIV+ women tended to have lower frequencies of CD56(Dim) NK cells in both blood (p = 0.0001) and warts (p = 0.006) than HIV- women. Wart NK cells from HIV+ women expressed significantly lower CD107a and produced IFN-γ. HAART status was not associated with differences in NK cell functionality. We conclude that wart NK cells from HIV+ women have defects in their ability to degranulate and/or secrete IFN-γ, which may provide insights into why HIV+ women fail to spontaneously resolve genital warts. Copyright © 2013 Elsevier Inc. All rights reserved.

The Effect of Disinfection on Viability and Function of Baboon Red Blood Cells and Platelets

DTIC Science & Technology

1997-07-11

blood cells was evaluated by their ability to transport oxygen as assessed by measurement of 2,3 diphosphoglycerate (DPG)14 and red blood cell p50,15...Blood collected from the bleeding time site (referred to as "shed blood") had a significantly reduced thromboxane A2 level . The ability of the...preserved or treated platelets to increase the shed blood thromboxane A2 level and reduce the 8; extended bleeding time is the measure of their

Impairment of T-regulatory cells in cord blood of atopic mothers.

PubMed

Schaub, Bianca; Liu, Jing; Höppler, Sabine; Haug, Severine; Sattler, Christine; Lluis, Anna; Illi, Sabina; von Mutius, Erika

2008-06-01

Maternal atopy is a strong predictor for the development of childhood allergic diseases. The underlying mechanisms are ill defined, yet regulatory T (Treg) and T(H)17 cells may play a key role potentially shaping the early immune system toward a proallergic or antiallergic immune regulation. We examined T(H)1/T(H)2, Treg, and T(H)17 cell responses to innate (lipid A/peptidoglycan) and mitogen/adaptive (phytohemagglutinin/Dermatophagoides pteronyssinus 1) immune stimulation in cord blood from offspring of atopic/nonatopic mothers. Cord blood mononuclear cells from 161 healthy neonates (59% nonatopic, 41% atopic mothers) were investigated regarding Treg and T(H)17 cells (mRNA/surface markers), suppressive function, and proliferation/cytokine secretion. Cord blood from offspring of atopic mothers showed fewer innate-induced Treg cells (CD4(+)CD25(+)high), lower mRNA expression of associated markers (glucocorticoid-induced tumor necrosis factor receptor-related protein/lymphocyte activation gene 3; P < .05), and a trend toward lower Forkhead box transcription factor 3 (Foxp3) expression. Treg cell function was impaired in mitogen-induced suppression of T effector cells in cord blood of offspring from atopic mothers (P = .03). Furthermore, IL-10 and IFN-gamma secretion were decreased in innate-stimulated cord blood of offspring from atopic mothers (P = .04/.05). Innate-induced IL-17 was independent of maternal atopy and highly correlated with IL-13 secretion. In offspring of atopic mothers, Treg cell numbers, expression, and function were impaired at birth. T(H)17 cells were correlated with T(H)2 cells, independently of maternal atopy.

Cryopreservation of red blood cells.

PubMed

Lagerberg, Johan W

2015-01-01

Cryopreservation of red blood cell concentrates (RBCs) is an important method for maintaining an inventory of rare RBC units and managing special transfusion circumstances. The permeating additive glycerol is used as a cryoprotectant to protect RBCs against freezing damage. The use of thawed RBCs was hampered a 24-h outdating period due to potential bacterial contamination when a functionally open system was used for addition and removal of the glycerol. With the introduction of a functionally closed system for the glycerolization and deglycerolization of RBC units, extended post-thaw storage became possible. Here, we describe the cryopreservation of red blood cells according to the high-glycerol method, using a functionally closed processing system.

On-Orbit, Immuno-Based, Label-Free White Blood Cell Counting System with Microelectromechanical Sensor Technology (OILWBCS-MEMS)

NASA Technical Reports Server (NTRS)

Edmonds, Jessica

2015-01-01

Aurora Flight Sciences, in partnership with Draper Laboratory, has developed a miniaturized system to count white blood cells in microgravity environments. The system uses MEMS technology to simultaneously count total white blood cells, the five white blood cell differential subgroups, and various lymphocyte subtypes. The OILWBCS-MEMS detection technology works by immobilizing an array of white blood cell-specific antibodies on small, gold-coated membranes. When blood flows across the membranes, specific cells' surface protein antigens bind to their corresponding antibodies. This binding can be measured and correlated to cell counts. In Phase I, the partners demonstrated surface chemistry sensitivity and specificity for total white blood cells and two lymphocyte subtypes. In Phase II, a functional prototype demonstrated end-to-end operation. This rugged, miniaturized device requires minimal blood sample preparation and will be useful for both space flight and terrestrial applications.

Functional characterization of novel ABCB6 mutations and their clinical implications in familial pseudohyperkalemia

PubMed Central

Andolfo, Immacolata; Russo, Roberta; Manna, Francesco; De Rosa, Gianluca; Gambale, Antonella; Zouwail, Soha; Detta, Nicola; Pardo, Catia Lo; Alper, Seth L.; Brugnara, Carlo; Sharma, Alok K.; De Franceschi, Lucia; Iolascon, Achille

2016-01-01

Isolated familial pseudohyperkalemia is a dominant red cell trait characterized by cold-induced ‘passive leak’ of red cell potassium ions into plasma. The causative gene of this condition is ABCB6, which encodes an erythrocyte membrane ABC transporter protein bearing the Langereis blood group antigen system. In this study analyzing three new families, we report the first functional characterization of ABCB6 mutants, including the homozygous mutation V454A, heterozygous mutation R276W, and compound heterozygous mutations R276W and R723Q (in trans). All these mutations are annotated in public databases, suggesting that familial pseudohyperkalemia could be common in the general population. Indeed, we identified variant R276W in one of 327 random blood donors (0.3%). Four weeks’ storage of heterozygous R276W blood cells resulted in massive loss of potassium compared to that from healthy control red blood cells. Moreover, measurement of cation flux demonstrated greater loss of potassium or rubidium ions from HEK-293 cells expressing ABCB6 mutants than from cells expressing wild-type ABCB6. The R276W/R723Q mutations elicited greater cellular potassium ion efflux than did the other mutants tested. In conclusion, ABCB6 missense mutations in red blood cells from subjects with familial pseudohyperkalemia show elevated potassium ion efflux. The prevalence of such individuals in the blood donor population is moderate. The fact that storage of blood from these subjects leads to significantly increased levels of potassium in the plasma could have serious clinical implications for neonates and infants receiving large-volume transfusions of whole blood. Genetic tests for familial pseudohyperkalemia could be added to blood donor pre-screening. Further study of ABCB6 function and trafficking could be informative for the study of other pathologies of red blood cell hydration. PMID:27151991

Generation of Functional Blood Vessels from a Single c-kit+ Adult Vascular Endothelial Stem Cell

PubMed Central

Fang, Shentong; Wei, Jing; Pentinmikko, Nalle; Leinonen, Hannele; Salven, Petri

2012-01-01

In adults, the growth of blood vessels, a process known as angiogenesis, is essential for organ growth and repair. In many disorders including cancer, angiogenesis becomes excessive. The cellular origin of new vascular endothelial cells (ECs) during blood vessel growth in angiogenic situations has remained unknown. Here, we provide evidence for adult vascular endothelial stem cells (VESCs) that reside in the blood vessel wall endothelium. VESCs constitute a small subpopulation within CD117+ (c-kit+) ECs capable of undergoing clonal expansion while other ECs have a very limited proliferative capacity. Isolated VESCs can produce tens of millions of endothelial daughter cells in vitro. A single transplanted c-kit-expressing VESC by the phenotype lin−CD31+CD105+Sca1+CD117+ can generate in vivo functional blood vessels that connect to host circulation. VESCs also have long-term self-renewal capacity, a defining functional property of adult stem cells. To provide functional verification on the role of c-kit in VESCs, we show that a genetic deficit in endothelial c-kit expression markedly decreases total colony-forming VESCs. In vivo, c-kit expression deficit resulted in impaired EC proliferation and angiogenesis and retardation of tumor growth. Isolated VESCs could be used in cell-based therapies for cardiovascular repair to restore tissue vascularization after ischemic events. VESCs also provide a novel cellular target to block pathological angiogenesis and cancer growth. PMID:23091420

An oncological view on the blood-testis barrier.

PubMed

Bart, Joost; Groen, Harry J M; van der Graaf, Winette T A; Hollema, Harry; Hendrikse, N Harry; Vaalburg, Willem; Sleijfer, Dirk T; de Vries, Elisabeth G E

2002-06-01

The function of the blood-testis barrier is to protect germ cells from harmful influences; thus, it also impedes the delivery of chemotherapeutic drugs to the testis. The barrier has three components: first, a physicochemical barrier consisting of continuous capillaries, Sertoli cells in the tubular wall, connected together with narrow tight junctions, and a myoid-cell layer around the seminiferous tubule. Second, an efflux-pump barrier that contains P-glycoprotein in the luminal capillary endothelium and on the myoid-cell layer; and multidrug-resistance associated protein 1 located basolaterally on Sertoli cells. Third, an immunological barrier, consisting of Fas ligand on Sertoli cells. Inhibition of P-glycoprotein function offers the opportunity to increase the delivery of cytotoxic drugs to the testis. In the future, visualisation of function in the blood-testis barrier may also be helpful to identify groups of patients in whom testis conservation is safe or to select drugs that are less harmful to fertility.

From artificial red blood cells, oxygen carriers, and oxygen therapeutics to artificial cells, nanomedicine, and beyond

PubMed Central

Chang, Thomas M. S.

2013-01-01

The first experimental artificial red blood cells have all three major functions of red blood cells (rbc). However, the first practical one is a simple polyhemoglobin (PolyHb) that only has an oxygen-carrying function. This is now in routine clinical use in South Africa and Russia. An oxygen carrier with antioxidant functions, PolyHb-catalase-superoxide dismutase, can fulfill two of the three functions of rbc. Even more complete is one with all three functions of rbc in the form of PolyHb-catalase-superoxide dismutase-carbonic anhydrase. The most advanced ones are nanodimension artificial rbc with either PEG-lipid membrane or PEG-PLA polymermembrane. Extensions in to oxygen therapeutics include a PolyHb-tyrosinase that suppresses the growth of melanoma in a mice model. Another is a PolyHb-fibrinogen that is an oxygen carrier with platelet-like function. Research has now extended well beyond the original research on artificial rbc into many areas of artificial cells. These include nanoparticles, nanotubules, lipid vesicles, liposomes, polymer-tethered lipid vesicles, polymersomes, microcapsules, bioencapsulation, nanocapules, macroencapsulation, synthetic cells, and others. These are being used in nanotechnology, nanomedicine, regenerative medicine, enzyme/gene therapy, cell/stem cell therapy, biotechnology, drug delivery, hemoperfusion, nanosensers, and even by some groups in agriculture, industry, aquatic culture, nanocomputers, and nanorobotics. PMID:22409281

Lin- CD34hi CD117int/hi FcεRI+ cells in human blood constitute a rare population of mast cell progenitors.

PubMed

Dahlin, Joakim S; Malinovschi, Andrei; Öhrvik, Helena; Sandelin, Martin; Janson, Christer; Alving, Kjell; Hallgren, Jenny

2016-01-28

Mast cells are rare tissue-resident immune cells that are involved in allergic reactions, and their numbers are increased in the lungs of asthmatics. Murine lung mast cells arise from committed bone marrow-derived progenitors that enter the blood circulation, migrate through the pulmonary endothelium, and mature in the tissue. In humans, mast cells can be cultured from multipotent CD34(+) progenitor cells. However, a population of distinct precursor cells that give rise to mast cells has remained undiscovered. To our knowledge, this is the first report of human lineage-negative (Lin(-)) CD34(hi) CD117(int/hi) FcεRI(+) progenitor cells, which represented only 0.0053% of the isolated blood cells in healthy individuals. These cells expressed integrin β7 and developed a mast cell-like phenotype, although with a slow cell division capacity in vitro. Isolated Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood cells had an immature mast cell-like appearance and expressed high levels of many mast cell-related genes as compared with human blood basophils in whole-transcriptome microarray analyses. Furthermore, serglycin, tryptase, and carboxypeptidase A messenger RNA transcripts were detected by quantitative reverse transcription-polymerase chain reaction. Altogether, we propose that the Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood cells are closely related to human tissue mast cells and likely constitute an immediate precursor population, which can give rise to predominantly mast cells. Furthermore, asthmatics with reduced lung function had a higher frequency of Lin(-) CD34(hi) CD117(int/hi) FcεRI(+) blood mast cell progenitors than asthmatics with normal lung function. © 2016 by The American Society of Hematology.

A novel perivascular cell population in the zebrafish brain

PubMed Central

Galanternik, Marina Venero; Castranova, Daniel; Gore, Aniket V; Blewett, Nathan H; Jung, Hyun Min; Stratman, Amber N; Kirby, Martha R; Iben, James; Miller, Mayumi F; Kawakami, Koichi; Maraia, Richard J; Weinstein, Brant M

2017-01-01

The blood-brain barrier is essential for the proper homeostasis and function of the CNS, but its mechanism of function is poorly understood. Perivascular cells surrounding brain blood vessels are thought to be important for blood-brain barrier establishment, but their roles are not well defined. Here, we describe a novel perivascular cell population closely associated with blood vessels on the zebrafish brain. Based on similarities in their morphology, location, and scavenger behavior, these cells appear to be the zebrafish equivalent of cells variably characterized as Fluorescent Granular Perithelial cells (FGPs), perivascular macrophages, or ‘Mato Cells’ in mammals. Despite their macrophage-like morphology and perivascular location, zebrafish FGPs appear molecularly most similar to lymphatic endothelium, and our imaging studies suggest that these cells emerge by differentiation from endothelium of the optic choroidal vascular plexus. Our findings provide the first report of a perivascular cell population in the brain derived from vascular endothelium. DOI: http://dx.doi.org/10.7554/eLife.24369.001 PMID:28395729

On the Mechanism of Human Red Blood Cell Longevity: Roles of Calcium, the Sodium Pump, PIEZO1, and Gardos Channels.

PubMed

Lew, Virgilio L; Tiffert, Teresa

2017-01-01

In a healthy adult, the transport of O 2 and CO 2 between lungs and tissues is performed by about 2 · 10 13 red blood cells, of which around 1.7 · 10 11 are renewed every day, a turnover resulting from an average circulatory lifespan of about 120 days. Cellular lifespan is the result of an evolutionary balance between the energy costs of maintaining cells in a fit functional state versus cell renewal. In this Review we examine how the set of passive and active membrane transporters of the mature red blood cells interact to maximize their circulatory longevity thus minimizing costs on expensive cell turnover. Red blood cell deformability is critical for optimal rheology and gas exchange functionality during capillary flow, best fulfilled when the volume of each human red blood cell is kept at a fraction of about 0.55-0.60 of the maximal spherical volume allowed by its membrane area, the optimal-volume-ratio range. The extent to which red blood cell volumes can be preserved within or near these narrow optimal-volume-ratio margins determines the potential for circulatory longevity. We show that the low cation permeability of red blood cells allows volume stability to be achieved with extraordinary cost-efficiency, favouring cell longevity over cell turnover. We suggest a mechanism by which the interplay of a declining sodium pump and two passive membrane transporters, the mechanosensitive PIEZO1 channel, a candidate mediator of P sickle in sickle cells, and the Ca 2+ -sensitive, K + -selective Gardos channel, can implement red blood cell volume stability around the optimal-volume-ratio range, as required for extended circulatory longevity.

On the Mechanism of Human Red Blood Cell Longevity: Roles of Calcium, the Sodium Pump, PIEZO1, and Gardos Channels

PubMed Central

Lew, Virgilio L.; Tiffert, Teresa

2017-01-01

In a healthy adult, the transport of O2 and CO2 between lungs and tissues is performed by about 2 · 1013 red blood cells, of which around 1.7 · 1011 are renewed every day, a turnover resulting from an average circulatory lifespan of about 120 days. Cellular lifespan is the result of an evolutionary balance between the energy costs of maintaining cells in a fit functional state versus cell renewal. In this Review we examine how the set of passive and active membrane transporters of the mature red blood cells interact to maximize their circulatory longevity thus minimizing costs on expensive cell turnover. Red blood cell deformability is critical for optimal rheology and gas exchange functionality during capillary flow, best fulfilled when the volume of each human red blood cell is kept at a fraction of about 0.55–0.60 of the maximal spherical volume allowed by its membrane area, the optimal-volume-ratio range. The extent to which red blood cell volumes can be preserved within or near these narrow optimal-volume-ratio margins determines the potential for circulatory longevity. We show that the low cation permeability of red blood cells allows volume stability to be achieved with extraordinary cost-efficiency, favouring cell longevity over cell turnover. We suggest a mechanism by which the interplay of a declining sodium pump and two passive membrane transporters, the mechanosensitive PIEZO1 channel, a candidate mediator of Psickle in sickle cells, and the Ca2+-sensitive, K+-selective Gardos channel, can implement red blood cell volume stability around the optimal-volume-ratio range, as required for extended circulatory longevity. PMID:29311949

Mesenchymal stem cells derived from peripheral blood protects against ischemia.

PubMed

Ukai, Ryo; Honmou, Osamu; Harada, Kuniaki; Houkin, Kiyohiro; Hamada, Hirofumi; Kocsis, Jeffery D

2007-03-01

Intravenous delivery of mesenchymal stem cells (MSCs) prepared from bone marrow (BMSCs) reduces infarction volume and ameliorates functional deficits in a rat cerebral ischemia model. MSC-like multipotent precursor cells (PMSCs) have also been suggested to exist in peripheral blood. To test the hypothesis that treatment with PMSCs may have a therapeutic benefit in stroke, we compared the efficacy of systemic delivery of BMSCs and PMSCs. A permanent middle cerebral artery occlusion (MCAO) in rat was induced by intraluminal vascular occlusion with a microfilament. Rat BMSCs and PMSCs were prepared in culture and intravenously injected into the rats 6 h after MCAO. Lesion size was assessed at 6 h, and 1, 3, and 7 days using MR imaging and histology. The hemodynamic change of cerebral blood perfusion on stroke was assessed the same times using perfusion-weighted image (PWI). Functional outcome was assessed using the treadmill stress test. Both BMSCs and PMSCs treated groups had reduced lesion volume, improved regional cerebral blood flow, and functional improvement compared to the control group. The therapeutic benefits of both MSC-treated groups were similar. These data suggest that PMSCs derived from peripheral blood could be an important cell source of cell therapy for stroke.

Generation of mesenchymal stromal cells in the presence of platelet lysate: a phenotypic and functional comparison of umbilical cord blood- and bone marrow-derived progenitors

PubMed Central

Avanzini, Maria Antonietta; Bernardo, Maria Ester; Cometa, Angela Maria; Perotti, Cesare; Zaffaroni, Nadia; Novara, Francesca; Visai, Livia; Moretta, Antonia; Del Fante, Claudia; Villa, Raffaella; Ball, Lynne M.; Fibbe, Willem E.; Maccario, Rita; Locatelli, Franco

2009-01-01

Background Mesenchymal stromal cells are employed in various different clinical settings in order to modulate immune response. However, relatively little is known about the mechanisms responsible for their immunomodulatory effects, which could be influenced by both the cell source and culture conditions. Design and Methods We tested the ability of a 5% platelet lysate-supplemented medium to support isolation and ex vivo expansion of mesenchymal stromal cells from full-term umbilical-cord blood. We also investigated the biological/functional properties of umbilical cord blood mesenchymal stromal cells, in comparison with platelet lysate-expanded bone marrow mesenchymal stromal cells. Results The success rate of isolation of mesenchymal stromal cells from umbilical cord blood was in the order of 20%. These cells exhibited typical morphology, immunophenotype and differentiation capacity. Although they have a low clonogenic efficiency, umbilical cord blood mesenchymal stromal cells may possess high proliferative potential. The genetic stability of these cells from umbilical cord blood was demonstrated by a normal molecular karyotype; in addition, these cells do not express hTERT and telomerase activity, do express p16ink4a protein and do not show anchorage-independent cell growth. Concerning alloantigen-specific immune responses, umbilical cord blood mesenchymal stromal cells were able to: (i) suppress T- and NK-lymphocyte proliferation, (ii) decrease cytotoxic activity and (iii) only slightly increase interleukin-10, while decreasing interferon-γ secretion, in mixed lymphocyte culture supernatants. While an indoleamine 2,3-dioxygenase-specific inhibitor did not reverse mesenchymal stromal cell-induced suppressive effects, a prostaglandin E2-specific inhibitor hampered the suppressive effect of both umbilical cord blood- and bone marrow-mesenchymal stromal cells on alloantigen-induced cytotoxic activity. Mesenchymal stromal cells from both sources expressed HLA-G. Conclusions Umbilical cord blood- and bone marrow-mesenchymal stromal cells may differ in terms of clonogenic efficiency, proliferative capacity and immunomodulatory properties; these differences may be relevant for clinical applications. PMID:19773264

Preparation and in vitro function of granulocyte concentrates for transfusion to neonates using the IBM 2991 blood processor.

PubMed

Goldfinger, D; Medici, M A; Hsi, R; McPherson, J; Connelly, M

1983-01-01

Clinical studies have suggested that granulocyte transfusions may be of value in the treatment of septic neonatal patients who present with severe granulocytopenia. We have developed a protocol for the preparation of granulocyte concentrates from freshly collected units of whole blood, using an automated blood cell processor. The red cells were washed with saline. Then, the buffy coats were collected from the washed red cells and studied for their suitability as granulocyte concentrates for neonatal transfusion. The mean number of granulocytes per concentrate was 1.6 X 10(9) in a mean volume of 25 ml. Studies of granulocyte function, including viability, random mobility, chemotaxis, phagocytosis and nitro-blue tetrazolium reduction, demonstrated that the granulocytes were functionally unimpaired following preparation of the concentrates. These studies suggest that concentrates of functional granulocytes, suitable for transfusion to neonatal patients, can be prepared from fresh units of whole blood, using a cell processor. This procedure is more cost-effective than leukapheresis and allows for delivery of granulocytes for transfusion in a more timely fashion.

Hearing Loss and Auditory Function in Sickle Cell Disease

ERIC Educational Resources Information Center

Burch-Sims, G.P.; Matlock, V.R.

2005-01-01

Sickle cell disease was first reported in 1910 by J. Herrick, and since then, various associated conditions and complications have been described. Sickle cell disease is a hereditary disorder characterized by abnormality of the hemoglobin in the red blood cell. During periods of decreased oxygen tension in the red blood cell's environment, the…

Blood cell interactions and segregation in flow.

PubMed

Munn, Lance L; Dupin, Michael M

2008-04-01

For more than a century, pioneering researchers have been using novel experimental and computational approaches to probe the mysteries of blood flow. Thanks to their efforts, we know that blood cells generally prefer to migrate to the axis of flow, that red and white cells segregate in flow, and that cell deformability and their tendency to reversibly aggregate contribute to the non-Newtonian nature of this unique fluid. All of these properties have beneficial physiological consequences, allowing blood to perform a variety of critical functions. Our current understanding of these unusual flow properties of blood have been made possible by the ingenuity and diligence of a number of researchers, including Harry Goldsmith, who developed novel technologies to visualize and quantify the flow of blood at the level of individual cells. Here we summarize efforts in our lab to continue this tradition and to further our understanding of how blood cells interact with each other and with the blood vessel wall.

In vitro fabrication of functional three-dimensional tissues with perfusable blood vessels

PubMed Central

Sekine, Hidekazu; Shimizu, Tatsuya; Sakaguchi, Katsuhisa; Dobashi, Izumi; Wada, Masanori; Yamato, Masayuki; Kobayashi, Eiji; Umezu, Mitsuo; Okano, Teruo

2013-01-01

In vitro fabrication of functional vascularized three-dimensional tissues has been a long-standing objective in the field of tissue engineering. Here we report a technique to engineer cardiac tissues with perfusable blood vessels in vitro. Using resected tissue with a connectable artery and vein as a vascular bed, we overlay triple-layer cardiac cell sheets produced from coculture with endothelial cells, and support the tissue construct with media perfused in a bioreactor. We show that endothelial cells connect to capillaries in the vascular bed and form tubular lumens, creating in vitro perfusable blood vessels in the cardiac cell sheets. Thicker engineered tissues can be produced in vitro by overlaying additional triple-layer cell sheets. The vascularized cardiac tissues beat and can be transplanted with blood vessel anastomoses. This technique may create new opportunities for in vitro tissue engineering and has potential therapeutic applications. PMID:23360990

IL-10+ regulatory B cells are enriched in cord blood and may protect against cGVHD after cord blood transplantation.

PubMed

Sarvaria, Anushruti; Basar, Rafet; Mehta, Rohtesh S; Shaim, Hila; Muftuoglu, Muharrem; Khoder, Ahmad; Sekine, Takuye; Gokdemir, Elif; Kondo, Kayo; Marin, David; Daher, May; Alousi, Amin M; Alsuliman, Abdullah; Liu, Enli; Oran, Betul; Olson, Amanda; Jones, Roy B; Popat, Uday; Hosing, Chitra; Champlin, Richard; Shpall, Elizabeth J; Rezvani, Katayoun

2016-09-08

Cord blood (CB) offers a number of advantages over other sources of hematopoietic stem cells, including a lower rate of chronic graft-versus-host disease (cGVHD) in the presence of increased HLA disparity. Recent research in experimental models of autoimmunity and in patients with autoimmune or alloimmune disorders has identified a functional group of interleukin-10 (IL-10)-producing regulatory B cells (Bregs) that negatively regulate T-cell immune responses. At present, however, there is no consensus on the phenotypic signature of Bregs, and their prevalence and functional characteristics in CB remain unclear. Here, we demonstrate that CB contains an abundance of B cells with immunoregulatory function. Bregs were identified in both the naive and transitional B-cell compartments and suppressed T-cell proliferation and effector function through IL-10 production as well as cell-to-cell contact involving CTLA-4. We further show that the suppressive capacity of CB-derived Bregs can be potentiated through CD40L signaling, suggesting that inflammatory environments may induce their function. Finally, there was robust recovery of IL-10-producing Bregs in patients after CB transplantation, to higher frequencies and absolute numbers than seen in the peripheral blood of healthy donors or in patients before transplant. The reconstituting Bregs showed strong in vitro suppressive activity against allogeneic CD4(+) T cells, but were deficient in patients with cGVHD. Together, these findings identify a rich source of Bregs and suggest a protective role for CB-derived Bregs against cGVHD development in CB recipients. This advance could propel the development of Breg-based strategies to prevent or ameliorate this posttransplant complication. © 2016 by The American Society of Hematology.

Human spleen and red blood cells

NASA Astrophysics Data System (ADS)

Pivkin, Igor; Peng, Zhangli; Karniadakis, George; Buffet, Pierre; Dao, Ming

2016-11-01

Spleen plays multiple roles in the human body. Among them is removal of old and altered red blood cells (RBCs), which is done by filtering cells through the endothelial slits, small micron-sized openings. There is currently no experimental technique available that allows us to observe RBC passage through the slits. It was previously noticed that people without a spleen have less deformable red blood cells, indicating that the spleen may play a role in defining the size and shape of red blood cells. We used detailed RBC model implemented within the Dissipative Particle Dynamics (DPD) simulation framework to study the filter function of the spleen. Our results demonstrate that spleen indeed plays major role in defining the size and shape of the healthy human red blood cells.

CD4+ T Cell Activation and Vascular Normalization: Two Sides of the Same Coin?

PubMed

De Palma, Michele; Jain, Rakesh K

2017-05-16

Normalization of tumor blood vessels enhances the infiltration and functions of T cells. Tian et al. (2017) report that effector CD4 + T cells, in turn, support vascular normalization, highlighting intertwined roles for blood vessels and T cells in cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

Hematological alterations and thymic function in newborns of HIV-infected mothers receiving antiretroviral drugs.

PubMed

Wongnoi, Rotjanee; Penvieng, Nawaporn; Singboottra, Panthong; Kingkeow, Doungnapa; Oberdorfer, Peninnah; Sirivatanapa, Pannee; Pornprasert, Sakorn

2013-06-08

To investigate the effects of antiretroviral (ARV) drugs on hematological parameters and thymic function in HIV-uninfected newborns of HIV-infected mothers. Cross sectional study. Chiang-Mai University Hospital, Chiang-Mai, Thailand. 49 HIV-uninfected and 26 HIV-infected pregnancies. Cord blood samples of newborns from HIV-uninfected and HIV-infected mothers were collected. Hematological parameters were measured using automatic blood cell count. T-cell receptor excision circles (TRECs) levels in cord blood mononuclear cells (CBMCs), CD4+ and CD8+ T-cells were quantified using real-time PCR.. Hemotological parameters and thymic function. Newborn of HIV-infected mother tended to have lower mean levels of hemoglobin than those of HIV-uninfected mother (137 ±22 vs 146 ±17 g/L, P = 0.05). Furthermore, mean of red blood cell (RBC) counts and hematocrit and median of TRECs in CD4+ T-cells in the newborns of the former were significantly lower than those of the latter [3.6 ±0.7 vs 4.8 ±0.6 x 1012 cells/L, P <0.001; 0.40 ±0.07 vs 0.46 ±0.05 L/L, P < 0.001 and 0.53 (IQR: 0.03-5.76) vs 13.20 (IQR: 2.77-27.51) x 10-3 pg/uL, P = 0.02, respectively]. ARV drugs altered hematological parameters and thymic function (TRECs CD4+ T-cells) in HIV-uninfected newborns of HIV-infected mothers.

Limited immune surveillance in lymphoid tissue by cytolytic CD4+ T cells during health and HIV disease

PubMed Central

McLane, Laura M.; Steblyanko, Maria; Anikeeva, Nadia; Ablanedo-Terrazas, Yuria; Demers, Korey; Eller, Michael A.; Streeck, Hendrik; Jansson, Marianne; Sönnerborg, Anders; Canaday, David H.; Naji, Ali; Wherry, E. John; Robb, Merlin L.; Reyes-Teran, Gustavo; Sykulev, Yuri; Betts, Michael R.

2018-01-01

CD4+ T cells subsets have a wide range of important helper and regulatory functions in the immune system. Several studies have specifically suggested that circulating effector CD4+ T cells may play a direct role in control of HIV replication through cytolytic activity or autocrine β-chemokine production. However, it remains unclear whether effector CD4+ T cells expressing cytolytic molecules and β-chemokines are present within lymph nodes (LNs), a major site of HIV replication. Here, we report that expression of β-chemokines and cytolytic molecules are enriched within a CD4+ T cell population with high levels of the T-box transcription factors T-bet and eomesodermin (Eomes). This effector population is predominately found in peripheral blood and is limited in LNs regardless of HIV infection or treatment status. As a result, CD4+ T cells generally lack effector functions in LNs, including cytolytic capacity and IFNγ and β-chemokine expression, even in HIV elite controllers and during acute/early HIV infection. While we do find the presence of degranulating CD4+ T cells in LNs, these cells do not bear functional or transcriptional effector T cell properties and are inherently poor to form stable immunological synapses compared to their peripheral blood counterparts. We demonstrate that CD4+ T cell cytolytic function, phenotype, and programming in the peripheral blood is dissociated from those characteristics found in lymphoid tissues. Together, these data challenge our current models based on blood and suggest spatially and temporally dissociated mechanisms of viral control in lymphoid tissues. PMID:29652923

In vivo acoustic and photoacoustic focusing of circulating cells

NASA Astrophysics Data System (ADS)

Galanzha, Ekaterina I.; Viegas, Mark G.; Malinsky, Taras I.; Melerzanov, Alexander V.; Juratli, Mazen A.; Sarimollaoglu, Mustafa; Nedosekin, Dmitry A.; Zharov, Vladimir P.

2016-03-01

In vivo flow cytometry using vessels as natural tubes with native cell flows has revolutionized the study of rare circulating tumor cells in a complex blood background. However, the presence of many blood cells in the detection volume makes it difficult to count each cell in this volume. We introduce method for manipulation of circulating cells in vivo with the use of gradient acoustic forces induced by ultrasound and photoacoustic waves. In a murine model, we demonstrated cell trapping, redirecting and focusing in blood and lymph flow into a tight stream, noninvasive wall-free transportation of blood, and the potential for photoacoustic detection of sickle cells without labeling and of leukocytes targeted by functionalized nanoparticles. Integration of cell focusing with intravital imaging methods may provide a versatile biological tool for single-cell analysis in circulation, with a focus on in vivo needleless blood tests, and preclinical studies of human diseases in animal models.

In vivo acoustic and photoacoustic focusing of circulating cells

PubMed Central

Galanzha, Ekaterina I.; Viegas, Mark G.; Malinsky, Taras I.; Melerzanov, Alexander V.; Juratli, Mazen A.; Sarimollaoglu, Mustafa; Nedosekin, Dmitry A.; Zharov, Vladimir P.

2016-01-01

In vivo flow cytometry using vessels as natural tubes with native cell flows has revolutionized the study of rare circulating tumor cells in a complex blood background. However, the presence of many blood cells in the detection volume makes it difficult to count each cell in this volume. We introduce method for manipulation of circulating cells in vivo with the use of gradient acoustic forces induced by ultrasound and photoacoustic waves. In a murine model, we demonstrated cell trapping, redirecting and focusing in blood and lymph flow into a tight stream, noninvasive wall-free transportation of blood, and the potential for photoacoustic detection of sickle cells without labeling and of leukocytes targeted by functionalized nanoparticles. Integration of cell focusing with intravital imaging methods may provide a versatile biological tool for single-cell analysis in circulation, with a focus on in vivo needleless blood tests, and preclinical studies of human diseases in animal models. PMID:26979811

Impact of dialyzer membrane on apoptosis and function of polymorphonuclear cells and cytokine synthesis by peripheral blood mononuclear cells in hemodialysis patients.

PubMed

Andreoli, Maria C C; Dalboni, Maria A; Watanabe, Renato; Manfredi, Silvia R; Canziani, Maria E F; Kallás, Esper G; Sesso, Ricardo C; Draibe, Sergio A; Balakrishnan, Vaidyanathapuram S; Jaber, Bertrand L; Liangos, Orfeas; Cendoroglo, Miguel

2007-12-01

In an in vivo crossover trial, we compared a cellulosic with a synthetic dialyzer with respect to polymorphonuclear cells (PMN) function and apoptosis, cytokine serum levels and synthesis by peripheral blood mononuclear cells (PBMC), and complement activation. Twenty hemodialysis (HD) patients were assigned in alternate order to HD with cellulose acetate (CA) or polysulfone (PS) dialyzer. After 2 weeks, patients were crossed over to the second dialyzer and treated for another 2 weeks. Apoptosis was assessed by flow cytometry in freshly isolated PMN. Phagocytosis and production of peroxide by PMN were studied by flow cytometry in whole blood. PBMC were isolated from blood samples and incubated for 24 h with or without lipopolysaccharide (LPS). There was no impact of dialyzer biocompatibility on PMN apoptosis and function, cytokine synthesis by PBMC or on their serum levels, serum levels of C3a, and terminal complement complex (TCC). Nevertheless, after HD, serum levels of complement correlated negatively with PMN phagocytosis and peroxide production, and positively with PMN apoptosis and cytokine production by PBMC. Although the results did not show a dialyzer advantage on the immunologic parameters, complement activation may have modulated cell function and apoptosis after HD.

The modulatory role of cytokines IL-4 and IL-17 in the functional activity of phagocytes in diabetic pregnant women.

PubMed

Fagundes, Danny L G; França, Eduardo L; Gonzatti, Michelangelo B; Rugde, Marilza V C; Calderon, Iracema M P; Honorio-França, Adenilda C

2018-01-01

The study investigated the role of cytokines IL-4 and IL-17 in the modulation of the functional activity of mononuclear phagocytes in diabetic pregnant women with hyperglycemia. Sixty pregnant women were assigned to the following groups: nondiabetic (ND), mild gestational hyperglycemia (MGH), gestational diabetes mellitus (GDM), or type 2 diabetes mellitus (DM2). The functional activity of phagocytes from maternal blood, cord blood, and colostrum was assessed by determining their superoxide release, phagocytosis, microbicidal activity, and intracellular Ca 2+ release. Irrespective of glycemic status, colostrum and blood cells treated with IL-4 and IL-17 increased superoxide release in the presence of enteropathogenic Escherichia coli (EPEC). The highest phagocytosis rate was observed in cells from the DM2 group treated with IL-4. In all the groups, phagocytes from colostrum, maternal blood, and cord blood exhibited higher microbicidal activity against EPEC when treated with cytokines. IL-17 increased intracellular Ca 2+ release by colostrum phagocytes in diabetic groups. The results indicate that the IL-4 and IL-17 modulate the functional activity of phagocytes in the maternal blood, cord blood, and colostrum of diabetic mother. The natural immunity resulting from the interaction between the cells and cytokines tested may be an alternative procedure to improve the prognosis of maternal and newborn infections. © 2017 APMIS. Published by John Wiley & Sons Ltd.

Myeloid leukemia factor is a conserved regulator of RUNX transcription factor activity involved in hematopoiesis.

PubMed

Bras, Stéphanie; Martin-Lannerée, Séverine; Gobert, Vanessa; Augé, Benoît; Breig, Osman; Sanial, Matthieu; Yamaguchi, Masamitsu; Haenlin, Marc; Plessis, Anne; Waltzer, Lucas

2012-03-27

Defining the function of the genes that, like RUNX1, are deregulated in blood cell malignancies represents an important challenge. Myeloid leukemia factors (MLFs) constitute a poorly characterized family of conserved proteins whose founding member, MLF1, has been associated with acute myeloid leukemia in humans. To gain insight into the functions of this family, we investigated the role of the Drosophila MLF homolog during blood cell development. Here we report that mlf controls the homeostasis of the Drosophila hematopoietic system. Notably, mlf participates in a positive feedback loop to fine tune the activity of the RUNX transcription factor Lozenge (LZ) during development of the crystal cells, one of the two main blood cell lineages in Drosophila. At the molecular level, our data in cell cultures and in vivo strongly suggest that MLF controls the number of crystal cells by protecting LZ from degradation. Remarkably, it appears that the human MLF1 protein can substitute for MLF in the crystal cell lineage. In addition, MLF stabilizes the human oncogenic fusion protein RUNX1-ETO and is required for RUNX1-ETO-induced blood cell disorders in a Drosophila model of leukemia. Finally, using the human leukemic blood cell line Kasumi-1, we show that MLF1 depletion impairs RUNX1-ETO accumulation and reduces RUNX1-ETO-dependent proliferation. Thus, we propose that the regulation of RUNX protein levels is a conserved feature of MLF family members that could be critical for normal and pathological blood cell development.

Nanosphere-based one-step strategy for efficient and nondestructive detection of circulating tumor cells.

PubMed

Wu, Ling-Ling; Wen, Cong-Ying; Hu, Jiao; Tang, Man; Qi, Chu-Bo; Li, Na; Liu, Cui; Chen, Lan; Pang, Dai-Wen; Zhang, Zhi-Ling

2017-08-15

Detecting viable circulating tumor cells (CTCs) without disruption to their functions for in vitro culture and functional study could unravel the biology of metastasis and promote the development of personalized anti-tumor therapies. However, existing CTC detection approaches commonly include CTC isolation and subsequent destructive identification, which damages CTC viability and functions and generates substantial CTC loss. To address the challenge of efficiently detecting viable CTCs for functional study, we develop a nanosphere-based cell-friendly one-step strategy. Immunonanospheres with prominent magnetic/fluorescence properties and extraordinary stability in complex matrices enable simultaneous efficient magnetic capture and specific fluorescence labeling of tumor cells directly in whole blood. The collected cells with fluorescent tags can be reliably identified, free of the tedious and destructive manipulations from conventional CTC identification. Hence, as few as 5 tumor cells in ca. 1mL of whole blood can be efficiently detected via only 20min incubation, and this strategy also shows good reproducibility with the relative standard deviation (RSD) of 8.7%. Moreover, due to the time-saving and gentle processing and the minimum disruption of immunonanospheres to cells, 93.8±0.1% of detected tumor cells retain cell viability and proliferation ability with negligible changes of cell functions, capacitating functional study on cell migration, invasion and glucose uptake. Additionally, this strategy exhibits successful CTC detection in 10/10 peripheral blood samples of cancer patients. Therefore, this nanosphere-based cell-friendly one-step strategy enables viable CTC detection and further functional analyses, which will help to unravel tumor metastasis and guide treatment selection. Copyright © 2017 Elsevier B.V. All rights reserved.

Blood Cell Interactions and Segregation in Flow

PubMed Central

Munn, Lance L.; Dupin, Michael M.

2009-01-01

For more than a century, pioneering researchers have been using novel experimental and computational approaches to probe the mysteries of blood flow. Thanks to their efforts, we know that blood cells generally prefer to migrate to the axis of flow, that red and white cells segregate in flow, and that cell deformability and their tendency to reversibly aggregate contribute to the non-Newtonian nature of this unique fluid. All of these properties have beneficial physiological consequences, allowing blood to perform a variety of critical functions. Our current understanding of these unusual flow properties of blood have been made possible by the ingenuity and diligence of a number of researchers, including Harry Goldsmith, who developed novel technologies to visualize and quantify the flow of blood at the level of individual cells. Here we summarize efforts in our lab to continue this tradition and to further our understanding of how blood cells interact with each other and with the blood vessel wall. PMID:18188702

Reprogramming of Adult Peripheral Blood Cells into Human Induced Pluripotent Stem Cells as a Safe and Accessible Source of Endothelial Cells.

PubMed

Simara, Pavel; Tesarova, Lenka; Rehakova, Daniela; Farkas, Simon; Salingova, Barbara; Kutalkova, Katerina; Vavreckova, Eva; Matula, Pavel; Matula, Petr; Veverkova, Lenka; Koutna, Irena

2018-01-01

New approaches in regenerative medicine and vasculogenesis have generated a demand for sufficient numbers of human endothelial cells (ECs). ECs and their progenitors reside on the interior surface of blood and lymphatic vessels or circulate in peripheral blood; however, their numbers are limited, and they are difficult to expand after isolation. Recent advances in human induced pluripotent stem cell (hiPSC) research have opened possible avenues to generate unlimited numbers of ECs from easily accessible cell sources, such as the peripheral blood. In this study, we reprogrammed peripheral blood mononuclear cells, human umbilical vein endothelial cells (HUVECs), and human saphenous vein endothelial cells (HSVECs) into hiPSCs and differentiated them into ECs. The phenotype profiles, functionality, and genome stability of all hiPSC-derived ECs were assessed and compared with HUVECs and HSVECs. hiPSC-derived ECs resembled their natural EC counterparts, as shown by the expression of the endothelial surface markers CD31 and CD144 and the results of the functional analysis. Higher expression of endothelial progenitor markers CD34 and kinase insert domain receptor (KDR) was measured in hiPSC-derived ECs. An analysis of phosphorylated histone H2AX (γH2AX) foci revealed that an increased number of DNA double-strand breaks upon reprogramming into pluripotent cells. However, differentiation into ECs restored a normal number of γH2AX foci. Our hiPSCs retained a normal karyotype, with the exception of the HSVEC-derived hiPSC line, which displayed mosaicism due to a gain of chromosome 1. Peripheral blood from adult donors is a suitable source for the unlimited production of patient-specific ECs through the hiPSC interstage. hiPSC-derived ECs are fully functional and comparable to natural ECs. The protocol is eligible for clinical applications in regenerative medicine, if the genomic stability of the pluripotent cell stage is closely monitored.

OXYGEN TRANSPORT IN THE MICROCIRCULATION AND ITS REGULATION

PubMed Central

Pittman, Roland N.

2012-01-01

Cells require energy to carry out their functions and they typically use oxidative phosphorylation to generate the needed ATP. Thus, cells have a continuous need for oxygen which they receive by diffusion from the blood through the interstitial fluid. The circulatory system pumps oxygen-rich blood through a network of increasingly minute vessels, the microcirculation. The structure of the microcirculation is such that all cells have at least one nearby capillary for diffusive exchange of oxygen and red blood cells release the oxygen bound to hemoglobin as they traverse capillaries. This review focuses first on the historical development of techniques to measure oxygen at various sites in the microcirculation, including the blood, interstitium and cells. Next, approaches are described as to how these techniques have been employed to make discoveries about different aspects of oxygen transport. Finally, ways in which oxygen might participate in the regulation of blood flow toward matching oxygen supply to oxygen demand is discussed. Overall, the transport of oxygen to the cells of the body is one of the most critical functions of the cardiovascular system and it is in the microcirculation where the final local determinants of oxygen supply, oxygen demand and their regulation are decided. PMID:23025284

Effect of medicinal mushrooms on blood cells under conditions of diabetes mellitus

PubMed Central

Vitak, Taras; Yurkiv, Borys; Wasser, Solomon; Nevo, Eviatar; Sybirna, Natalia

2017-01-01

Diabetes mellitus (DM) is the third most common non-infectious disease leading to early disability and high mortality. Moreover, the number of patients is growing every year. The main symptom of DM is hyperglycemia. Increased levels of blood glucose activate polyol, hexosamine, and protein kinase metabolic pathways cause the intensification of non-enzymatic glycosylation and nitration of macromolecules. This, in turn, leads to the development of oxidative and nitrative stresses and secondary complications, such as different kinds of micro- and macroangiopathies. Metabolic disorders caused by insulin deficiency in diabetes significantly impede the functioning of a homeostasis system, which change the physical, biochemical, morphological, and functional properties of blood cells. As a result, the oxygen-transport function of red blood cells (RBCs), rheological properties of the blood, and functions of immunocompetent cells as well as the process of apoptosis are primarily affected. Modern pharmacotherapy focuses on the search for new preparations that aim to decrease blood glucose levels. Undesirable side effects and adverse reactions caused by synthetic medicines led to the search and investigation of new preparations of natural origin. Medicinal mushrooms play an important role among such new preparations. They are a source of a large number of high- and low-molecular compounds with pronounced biological effects. Our investigations show pronounced hypoglycemic and anti-anemic action of submerged cultivated mycelium powder of medicinal mushrooms Agaricus brasiliensis (A. brasiliensis) and Ganoderma lucidum (G. lucidum) on streptozotocin-induced DM in rats. Also, we showed that mycelium powders have membrane protective properties as evidenced by the redistribution of RBC populations towards the growth of full functional cell numbers. Normalization of parameters of leukocyte formula and suppression of apoptosis of white blood cells in diabetic rats treated with A. brasiliensis and G. lucidum mycelia indicates pronounced positive effects of these strains of mushrooms. Thus, the use of medicinal mushrooms for treatment of DM and in prevention development of its secondary complications might be a new effective approach of this disease’s cure. This article is aimed at summarizing and analyzing the literature data and basic achievements concerning DM type 1 treatment using medicinal mushrooms and showing the results obtained in our research. PMID:28572880

Effect of medicinal mushrooms on blood cells under conditions of diabetes mellitus.

PubMed

Vitak, Taras; Yurkiv, Borys; Wasser, Solomon; Nevo, Eviatar; Sybirna, Natalia

2017-05-15

Diabetes mellitus (DM) is the third most common non-infectious disease leading to early disability and high mortality. Moreover, the number of patients is growing every year. The main symptom of DM is hyperglycemia. Increased levels of blood glucose activate polyol, hexosamine, and protein kinase metabolic pathways cause the intensification of non-enzymatic glycosylation and nitration of macromolecules. This, in turn, leads to the development of oxidative and nitrative stresses and secondary complications, such as different kinds of micro- and macroangiopathies. Metabolic disorders caused by insulin deficiency in diabetes significantly impede the functioning of a homeostasis system, which change the physical, biochemical, morphological, and functional properties of blood cells. As a result, the oxygen-transport function of red blood cells (RBCs), rheological properties of the blood, and functions of immunocompetent cells as well as the process of apoptosis are primarily affected. Modern pharmacotherapy focuses on the search for new preparations that aim to decrease blood glucose levels. Undesirable side effects and adverse reactions caused by synthetic medicines led to the search and investigation of new preparations of natural origin. Medicinal mushrooms play an important role among such new preparations. They are a source of a large number of high- and low-molecular compounds with pronounced biological effects. Our investigations show pronounced hypoglycemic and anti-anemic action of submerged cultivated mycelium powder of medicinal mushrooms Agaricus brasiliensis ( A. brasiliensis ) and Ganoderma lucidum ( G. lucidum ) on streptozotocin-induced DM in rats. Also, we showed that mycelium powders have membrane protective properties as evidenced by the redistribution of RBC populations towards the growth of full functional cell numbers. Normalization of parameters of leukocyte formula and suppression of apoptosis of white blood cells in diabetic rats treated with A. brasiliensis and G. lucidum mycelia indicates pronounced positive effects of these strains of mushrooms. Thus, the use of medicinal mushrooms for treatment of DM and in prevention development of its secondary complications might be a new effective approach of this disease's cure. This article is aimed at summarizing and analyzing the literature data and basic achievements concerning DM type 1 treatment using medicinal mushrooms and showing the results obtained in our research.

Differentiation-associated alteration in human monocyte-macrophage accessory cell function.

PubMed

Mayernik, D G; Ul-Haq, A; Rinehart, J J

1983-05-01

Human monocyte (Mo) to macrophage (Mx) differentiation is associated with marked and well studied changes in morphology, biochemical parameters, and effector cell function. Nevertheless, the comparative accessory cell (AC) function of blood Mo and differentiated Mx has not been carefully studied. We, therefore, examined the kinetics and mechanisms of change in AC function during in vitro Mo to Mx differentiation. The system utilized has two distinctive features: blood Mo and resultant cultured Mx represent a cohort of cells derived from the bone marrow within a 12-hr period. Moreover, the in vitro derived Mx utilized herein have been characterized extensively and are functionally and biochemically similar to pulmonary macrophages (PMx). In the experiments reported, AC functions of blood Mo, Mx derived from Mo after 1 to 6 days of culture, and PMx was compared. AC were cultured with nylon wool column-purified autologous T cells and were stimulated with concanavalin A (Con A) or streptokinase-streptodornase (SKSD). Blood T cell proliferation to Con A or SKSD was inhibited greater than 90% by the removal of Mo and was reconstituted by 20% Mo. Mx derived from Mo by culture for 1 to 3 days exhibited the same (or better) AC function as Mo when T cells were stimulated with either SKSD or Con A. In marked contrast, Mx derived from 6-day cultures exhibited less than or equal to 15% of Mo (i.e., control) capacity to support T cell proliferative response to SKSD. Six-day Mx support T cell proliferation to Con A was somewhat variable. Similar to 6-day cultured Mx, PMx failed to function as AC. The mechanism of loss of AC function was examined: a) cultured Mx maintained Ia antigen positivity for greater than 8 days; b) mixing experiments with Mo + 6-day cultured Mx or Mo + PMx demonstrated no T cell suppression; c) the normal capacity of most 6-day cultured Mx to support Con A but not SKSD induced T cell proliferation, apparently ruled out the loss of the ability to deliver a nonspecific "second signal" as the involved mechanism; d) inhibition of Mo to Mx differentiation by dexamethasone preserved AC activity. Thus, human culture-derived Mx and PMx exhibit deficit AC function through loss of an undefined mechanism. However, loss of AC antigen processing or presentation may occur.

[The specific features of the blood gas transport system in patients with postinfarction cardiosclerosis].

PubMed

Mikashinovich, Z I; Suroedova, R A; Olempieva, E V

2009-10-01

The specific features of blood gas transport system functioning were analyzed in patients with cardiovascular diseases. In patients with postinfarction cardiosclerosis (PICS), the quantitative mechanism for hypoxia adaptation tended to decrease, which may be considered to be a compensatory-adaptive reaction aimed at eliminating the sludge phenomenon and improving the rheological characteristics of blood. Acute myocardial reinfarction developed in patents with PICS is characterized by the lower functional activity of red blood cells, and developing hypoxia is an important link of activation of apoptotic cell death. The degree of hypoxia may be believed to correlate with the sizes of a myocardial necrosis focus.

Tissue-specific expression of transgenic secreted ACE in vasculature can restore normal kidney functions, but not blood pressure, of Ace-/- mice.

PubMed

Chattopadhyay, Saurabh; Kessler, Sean P; Colucci, Juliana Almada; Yamashita, Michifumi; Senanayake, Preenie deS; Sen, Ganes C

2014-01-01

Angiotensin-converting enzyme (ACE) regulates normal blood pressure and fluid homeostasis through its action in the renin-angiotensin-system (RAS). Ace-/- mice are smaller in size, have low blood pressure and defective kidney structure and functions. All of these defects are cured by transgenic expression of somatic ACE (sACE) in vascular endothelial cells of Ace-/- mice. sACE is expressed on the surface of vascular endothelial cells and undergoes a natural cleavage secretion process to generate a soluble form in the body fluids. Both the tissue-bound and the soluble forms of ACE are enzymatically active, and generate the vasoactive octapeptide Angiotensin II (Ang II) with equal efficiency. To assess the relative physiological roles of the secreted and the cell-bound forms of ACE, we expressed, in the vascular endothelial cells of Ace-/- mice, the ectodomain of sACE, which corresponded to only the secreted form of ACE. Our results demonstrated that the secreted form of ACE could normalize kidney functions and RAS integrity, growth and development of Ace-/- mice, but not their blood pressure. This study clearly demonstrates that the secreted form of ACE cannot replace the tissue-bound ACE for maintaining normal blood pressure; a suitable balance between the tissue-bound and the soluble forms of ACE is essential for maintaining all physiological functions of ACE.

Tissue-Specific Expression of Transgenic Secreted ACE in Vasculature Can Restore Normal Kidney Functions, but Not Blood Pressure, of Ace-/- Mice

PubMed Central

Chattopadhyay, Saurabh; Kessler, Sean P.; Colucci, Juliana Almada; Yamashita, Michifumi; Senanayake, Preenie deS; Sen, Ganes C.

2014-01-01

Angiotensin-converting enzyme (ACE) regulates normal blood pressure and fluid homeostasis through its action in the renin-angiotensin-system (RAS). Ace-/- mice are smaller in size, have low blood pressure and defective kidney structure and functions. All of these defects are cured by transgenic expression of somatic ACE (sACE) in vascular endothelial cells of Ace-/- mice. sACE is expressed on the surface of vascular endothelial cells and undergoes a natural cleavage secretion process to generate a soluble form in the body fluids. Both the tissue-bound and the soluble forms of ACE are enzymatically active, and generate the vasoactive octapeptide Angiotensin II (Ang II) with equal efficiency. To assess the relative physiological roles of the secreted and the cell-bound forms of ACE, we expressed, in the vascular endothelial cells of Ace-/- mice, the ectodomain of sACE, which corresponded to only the secreted form of ACE. Our results demonstrated that the secreted form of ACE could normalize kidney functions and RAS integrity, growth and development of Ace-/- mice, but not their blood pressure. This study clearly demonstrates that the secreted form of ACE cannot replace the tissue-bound ACE for maintaining normal blood pressure; a suitable balance between the tissue-bound and the soluble forms of ACE is essential for maintaining all physiological functions of ACE. PMID:24475296

Dragon's Blood Sap (Croton Lechleri) As Storage Medium For Avulsed Teeth: In Vitro Study Of Cell Viability.

PubMed

Martins, Christine Men; Hamanaka, Elizane Ferreira; Hoshida, Thayse Yumi; Sell, Ana Maria; Hidalgo, Mirian Marubayashi; Silveira, Catarina Soares; Poi, Wilson Roberto

2016-01-01

Tooth replantation success depends on the condition of cementum periodontal ligament after tooth avulsion; which is influenced by storage medium. The dragon's blood (Croton lechleri) sap has been suggested as a promising medium because it supports collagen formation and exhibits healing, anti-inflammatory and antimicrobial properties. Thus, the aim of this study was to evaluate the efficacy of dragon's blood sap as a storage medium for avulsed teeth through evaluation of functional and metabolic cell viability. This in vitro study compared the efficacy of different storage media to maintain the viability of human peripheral blood mononuclear and periodontal ligament cells. A 10% dragon's blood sap was tested while PBS was selected as its control. Ultra pasteurized whole milk was used for comparison as a commonly used storage medium. DMEM and distilled water were the positive and negative controls, respectively. The viability was assessed through trypan blue exclusion test and colorimetric MTT assay after 1, 3, 6, 10 and 24 h of incubation. The dragon's blood sap showed promising results due to its considerable maintenance of cell viability. For trypan blue test, the dragon's blood sap was similar to milk (p<0.05) and both presented the highest viability values. For MTT, the dragon's blood sap showed better results than all storage media, even better than milk (p<0.05). It was concluded that the dragon's blood sap was as effective as milk, the gold standard for storage medium. The experimental sap preserved the membrane of all cells and the functional viability of periodontal ligament cells.

Basic Components of Vascular Connective Tissue and Extracellular Matrix.

PubMed

Halper, Jaroslava

2018-01-01

Though the composition of the three layers constituting the blood vessel wall varies among the different types of blood vessels, and some layers may even be missing in capillaries, certain basic components, and properties are shared by all blood vessels, though each histologically distinct layer contains a unique complement of extracellular components, growth factors and cytokines, and cell types as well. The structure and composition of vessel layers informs and is informed by the function of the particular blood vessel. The adaptation of the composition and the resulting function of the extracellular matrix (ECM) to changes in circulation/blood flow and a variety of other extravascular stimuli can be characterized as remodeling spearheaded by vascular cells. There is a surprising amount of cell traffic among the three layers. It starts with endothelial cell mediated transmigration of inflammatory cells from the bloodstream into the subendothelium, and then into tissue adjoining the blood vessel. Smooth muscle cells and a variety of adventitial cells reside in tunica media and tunica externa, respectively. The latter cells are a mixture of progenitor/stem cells, fibroblasts, myofibroblasts, pericytes, macrophages, and dendritic cells and respond to endothelial injury by transdifferentiation as they travel into the two inner layers, intima and media for corrective mission in the ECM composition. This chapter addresses the role of various vascular cell types and ECM components synthesized by them in maintenance of normal structure and in their contribution to major pathological processes, such as atherosclerosis, organ fibrosis, and diabetic retinopathy. © 2018 Elsevier Inc. All rights reserved.

Reverse-D-4F Increases the Number of Endothelial Progenitor Cells and Improves Endothelial Progenitor Cell Dysfunctions in High Fat Diet Mice.

PubMed

Nana, Yang; Peng, Jiao; Jianlin, Zhang; Xiangjian, Zhang; Shutong, Yao; Enxin, Zhan; Bin, Li; Chuanlong, Zong; Hua, Tian; Yanhong, Si; Yunsai, Du; Shucun, Qin; Hui, Wang

2015-01-01

Although high density lipoprotein (HDL) improves the functions of endothelial progenitor cells (EPCs), the effect of HDL ApoAI mimetic peptide reverse-D-4F (Rev-D4F) on EPC mobilization and repair of EPC dysfunctions remains to be studied. In this study, we investigated the effects of Rev-D4F on peripheral blood cell subpopulations in C57 mice treated with a high fat diet and the mechanism of Rev-D4F in improving the function of EPCs impaired by tumor necrosis factor-α (TNF-α). The high fat diet significantly decreased the number of EPCs, EPC migratory functions, and the percentage of lymphocytes in the white blood cells. However, it significantly increased the number of white blood cells, the percentage of monocytes in the white blood cells, and the level of vascular endothelial growth factor (VEGF) and TNF-α in the plasma. Rev-D4F clearly inhibited the effect of the high fat diet on the quantification of peripheral blood cell subpopulations and cytokine levels, and increased stromal cell derived factor 1α (SDF-1α) in the plasma. We provided in vitro evidence that TNF-α impaired EPC proliferation, migration, and tube formation through inactive AKT and eNOS, which was restored by Rev-D4F treatment. In contrast, both the PI3-kinase (PI3K) inhibitor (LY294002) and AKT inhibitor (perifosine) obviously inhibited the restoration of Rev-4F on EPCs impaired by TNF-α. Our results suggested that Rev-D4F increases the quantity of endothelial progenitor cells through increasing the SDF-1α levels and decreasing the TNF-α level of peripheral blood in high fat diet-induced C57BL/6J mice, and restores TNF-α induced dysfunctions of EPCs partly through stimulating the PI3K/AKT signal pathway.

Induced Pluripotent Stem Cell-Derived Red Blood Cells and Platelet Concentrates: From Bench to Bedside.

PubMed

Focosi, Daniele; Amabile, Giovanni

2017-12-27

Red blood cells and platelets are anucleate blood components indispensable for oxygen delivery and hemostasis, respectively. Derivation of these blood elements from induced pluripotent stem (iPS) cells has the potential to develop blood donor-independent and genetic manipulation-prone products to complement or replace current transfusion banking, also minimizing the risk of alloimmunization. While the production of erythrocytes from iPS cells has challenges to overcome, such as differentiation into adult-type phenotype that functions properly after transfusion, platelet products are qualitatively and quantitatively approaching a clinically-applicable level owing to advances in expandable megakaryocyte (MK) lines, platelet-producing bioreactors, and novel reagents. Guidelines that assure the quality of iPS cells-derived blood products for clinical application represent a novel challenge for regulatory agencies. Considering the minimal risk of tumorigenicity and the expected significant demand of such products, ex vivo production of iPS-derived blood components can pave the way for iPS translation into the clinic.

Myeloid leukemia factor is a conserved regulator of RUNX transcription factor activity involved in hematopoiesis

PubMed Central

Bras, Stéphanie; Martin-Lannerée, Séverine; Gobert, Vanessa; Augé, Benoît; Breig, Osman; Sanial, Matthieu; Yamaguchi, Masamitsu; Haenlin, Marc; Plessis, Anne; Waltzer, Lucas

2012-01-01

Defining the function of the genes that, like RUNX1, are deregulated in blood cell malignancies represents an important challenge. Myeloid leukemia factors (MLFs) constitute a poorly characterized family of conserved proteins whose founding member, MLF1, has been associated with acute myeloid leukemia in humans. To gain insight into the functions of this family, we investigated the role of the Drosophila MLF homolog during blood cell development. Here we report that mlf controls the homeostasis of the Drosophila hematopoietic system. Notably, mlf participates in a positive feedback loop to fine tune the activity of the RUNX transcription factor Lozenge (LZ) during development of the crystal cells, one of the two main blood cell lineages in Drosophila. At the molecular level, our data in cell cultures and in vivo strongly suggest that MLF controls the number of crystal cells by protecting LZ from degradation. Remarkably, it appears that the human MLF1 protein can substitute for MLF in the crystal cell lineage. In addition, MLF stabilizes the human oncogenic fusion protein RUNX1-ETO and is required for RUNX1-ETO–induced blood cell disorders in a Drosophila model of leukemia. Finally, using the human leukemic blood cell line Kasumi-1, we show that MLF1 depletion impairs RUNX1-ETO accumulation and reduces RUNX1-ETO–dependent proliferation. Thus, we propose that the regulation of RUNX protein levels is a conserved feature of MLF family members that could be critical for normal and pathological blood cell development. PMID:22411814

Computed aided system for separation and classification of the abnormal erythrocytes in human blood

NASA Astrophysics Data System (ADS)

Wąsowicz, Michał; Grochowski, Michał; Kulka, Marek; Mikołajczyk, Agnieszka; Ficek, Mateusz; Karpieńko, Katarzyna; Cićkiewicz, Maciej

2017-12-01

The human peripheral blood consists of cells (red cells, white cells, and platelets) suspended in plasma. In the following research the team assessed an influence of nanodiamond particles on blood elements over various periods of time. The material used in the study consisted of samples taken from ten healthy humans of various age, different blood types and both sexes. The markings were leaded by adding to the blood unmodified diamonds and oxidation modified. The blood was put under an impact of two diamond concentrations: 20μl and 100μl. The amount of abnormal cells increased with time. The percentage of echinocytes as a result of interaction with nanodiamonds in various time intervals for individual specimens was scarce. The impact of the two diamond types had no clinical importance on red blood cells. It is supposed that as a result of longlasting exposure a dehydratation of red cells takes place, because of the function of the cells. The analysis of an influence of nanodiamond particles on blood elements was supported by computer system designed for automatic counting and classification of the Red Blood Cells (RBC). The system utilizes advanced image processing methods for RBCs separation and counting and Eigenfaces method coupled with the neural networks for RBCs classification into normal and abnormal cells purposes.

Hemoglobin Function in Stored Blood.

DTIC Science & Technology

1974-08-01

States during 1973. Several advantages over ACA) are important. Blood stored in CPD maintains higher ./ levels of 2,3-DPG (2,3- diphosphoglycerate ) and a...survival and ATP levels in stored blood is explained by the several functions of ATP which are necessary for cell viability. However, ATP levels do...not correlate with oxygen affinity during storage. Levels of 2,3-DPG determine oxygen affinity and thus hemoglobin function. (12,13) When normal levels

Which side of the balance determines the frequency of vaso-occlusive crises in children with sickle cell anemia: Blood viscosity or microvascular dysfunction?

PubMed

Charlot, Keyne; Romana, Marc; Moeckesch, Berenike; Jumet, Stéphane; Waltz, Xavier; Divialle-Doumdo, Lydia; Hardy-Dessources, Marie-Dominique; Petras, Marie; Tressières, Benoît; Tarer, Vanessa; Hue, Olivier; Etienne-Julan, Maryse; Antoine-Jonville, Sophie; Connes, Philippe

2016-01-01

Vascular resistance and tissue perfusion may be both affected by impaired vascular function and increased blood viscosity. Little is known about the effects of vascular function on the occurrence of painful vaso-occlusive crises (VOC) in children with sickle cell anemia (SCA). The aim of the present study was to determine which side of the balance (blood viscosity or vascular function) is the most deleterious in SCA and increases the risk for frequent hospitalized VOC. Microvascular function, microcirculatory oxygenation and blood viscosity were determined in a group of 22 SCA children/adolescents at steady state and a group of 13 healthy children/adolescents. Univariate analyses demonstrated blunted microvascular reactivity during local thermal heating test and decreased microcirculatory oxygenation in SCA children compared to controls. Multivariate analysis revealed that increased blood viscosity and decreased microcirculatory oxygenation were independent risk factors of frequent VOC in SCA. In contrast, the level of microvascular dysfunction does not predict VOC rate. In conclusion, increased blood viscosity is usually well supported in healthy individuals where vascular function is not impaired. However, in the context of SCA, microvascular function is impaired and any increase of blood viscosity or decrease in microcirculatory oxygenation would increase the risks for frequent VOC. Copyright © 2015 Elsevier Inc. All rights reserved.

Chronic heart failure and aging - effects of exercise training on endothelial function and mechanisms of endothelial regeneration: Results from the Leipzig Exercise Intervention in Chronic heart failure and Aging (LEICA) study.

PubMed

Sandri, Marcus; Viehmann, Manuel; Adams, Volker; Rabald, Kristin; Mangner, Norman; Höllriegel, Robert; Lurz, Philipp; Erbs, Sandra; Linke, Axel; Kirsch, Katharina; Möbius-Winkler, Sven; Thiery, Joachim; Teupser, Daniel; Hambrecht, Rainer; Schuler, Gerhard; Gielen, Stephan

2016-03-01

A reduction in number and function of endothelial progenitor cells (EPCs) occurs in both physiologic aging and chronic heart failure (CHF). We assessed whether disease and aging have additive effects on EPCs or whether beneficial effects of exercise training are diminished in old age. We randomized 60 patients with stable CHF and 60 referent controls to a training or a control group. To detect possible aging effects we included subjects below 55 (young) and above 65 years (older). Subjects in the training group exercised four times daily at 60% to 70% of VO2max for four weeks under supervision. At baseline and after the intervention the number and function of EPCs were assessed. As compared with young referent controls, older referent controls showed at baseline a reduced EPC number (young: 190 ± 37 CD34/KDR positive cells/ml blood; older: 131 ± 26 CD34/KDR positive cells/ml blood; p < 0.05) and function (young: 230 ± 41 migrated cells/1000 plated cells; older: 185 ± 28 cells/1000 plated cells; p < 0.05). In young and older CHF patients EPC-number (young: 85 ± 21 CD34/KDR positive cells/ml blood; older: 78 ± 20 CD34/KDR positive cells/ml blood) and EPC-function (young: 113 ± 26 cells/1000 plated cells; older: 120 ± 27 cells/1000 plated cells) were impaired. As a result of exercise training, EPC function improved by 24% in older referent controls (p < 0.05), while it remained unchanged in young training referent controls and controls respectively. In young and older patients with CHF four weeks of exercise training resulted in a significant improvement in EPC numbers and EPC function (young: number +66% function +43%; p < 0.05; older: number +69% function +36%; p < 0.05). These results were accompanied by a significant increase in flow mediated dilatation in the training groups of young/older CHF patients and in older referent controls. Four weeks of exercise training are effective in improving EPC number and EPC function in CHF patients. These training effects were not impaired among older patients, emphasizing the potentials of rehabilitation interventions in a patient group where CHF has a high prevalence. © The European Society of Cardiology 2015.

Alteration of functional state of peripheral blood erythrocytes in women of different age groups at dislipidemia conditions.

PubMed

Ratiani, L; Intskirveli, N; Ormotsadze, G; Sanikidze, T

2011-12-01

The aim of the study was identification of statistically reliable correlations and the cause-effect relationships between viability of red blood cells and dislipidema parametres and/or metabolic disorders, induced by age related alterations of estrogen content, in women of different ages (reproductive, menopausal) On the basis of the analysis of research results we can conclude that in the different age groups of women with atherosclerosis-induced cardiovascular diseases revealed estrogen-related dependence between Tg-s and HDL content, functional status of phereperial blood erytrotcites and severity of dislipidemia. The aterogenic index Tg/HD proved to be sensitive marker of dislipidemia in reproductive aging women, but does't reflect disorders of lipid metabolism in postmenosal women. It was proved the existence of reliable corelation between red blood cells dysfunction indicator, spherulation quality, and atherogenic index Tg/HDL highlights; however, the correlation coefficient is 2 times higher in the reproductive age as in menopause. Spherulation quality of red blood cells at low HDL content showd fast growth rate in reproductive-aged women, and was unsensetive to HDL content in postmenopasal women. It was concluded that age-related lack of estrogens in postmenopausal women indirectly contributes to decrease protection of red blood cells against oxidative damage, reduces their deformabelity and disturbances the rheological properties. So, Spherulation quality of red blood cells may be used as a diagnostic marker of severity of atherosclerosis.

Strategy for the hemocompatibility testing of microparticles.

PubMed

Braune, S; Basu, S; Kratz, K; Johansson, J Bäckemo; Reinthaler, M; Lendlein, A; Jung, F

2016-01-01

Polymer-based microparticles are applied as non-thrombogenic or thrombogenic materials in a wide variety of intra- or extra-corporeal medical devices. As demanded by the regulatory agencies, the hemocompatibility of these blood contacting biomaterials has to be evaluated in vitro to ensure that the particle systems appropriately fulfill the envisioned function without causing undesired events such as thrombosis or inflammation. Currently described in vitro assays for hemocompatibility testing of particles comprise tests with different single cell types (e.g. erythrocytes or leukocytes), varying concentrations/dilutions of the used blood cells or whole blood, which are not standardized.Here, we report about an in vitro dynamic test system for studying the hemocompatibility of polymeric microparticles utilizing fresh human whole blood from apparently healthy subjects, collected and processed under standardized conditions. Spherical poly(ether imide) microparticles with an average diameter of 140±30 μm were utilized as model systems. Reported as candidate materials for the removal of uremic toxins, these microparticles are anticipated to facilitate optimal flow conditions in a dialyzer with minimal backflow and blood cell damage. Pristine (PEI) and potassium hydroxide (PEI-KOH) functionalized microparticles exhibited similarly nanoporous surfaces (PEI: ØExternal pore = 90±60 nm; PEI-KOH ØExternal pore = 150±130 nm) but varying water wettabilities (PEI: θadv = 112±10° PEI-KOH θadv = 60±2°). The nanoporosity of the microparticle surfaces allows the exchange of toxic solutes from blood towards the interconnective pores in the particle core, while an immigration of the substantially larger blood cells is inhibited.Sterilized PEI microparticles were incorporated -air-free -in a syringe-based test system and exposed to whole blood for 60 minutes under gentle agitation. Thereafter, thrombi formation on the particles surfaces were analyzed microscopically. In the collected whole blood the non-adherent/circulating single blood cells were quantified via a differentiated complete blood cell count and the activation of platelets (P-Selectin expression, secretion and release), platelet function (PFA100 closure time) as well as thrombin formation (thrombin-antithrombin-complex) was analyzed. Free hemoglobin (HGB) levels were quantified as a measure of hemolysis.Microscopic evaluation revealed thrombi formation and particle aggregates for all tested microparticles. Reduction of circulating blood cells differed significantly between the particle types. Particularly, platelet and monocyte counts decreased up to 50% compared to the control (syringe filled with whole blood but without microparticles). In accordance, platelet activation, thrombin levels and degrees of hemolysis were clearly elevated in the particle loaded test systems and allowed a differentiation between the particle types. Increased PFA100 closure times (as activating agent a combination of collagen/ADP was used) indicated a similarly reduced ability of platelets to adhere and form stable aggregates independent from the particle type tested. This observation is most probably a consequence of the strong thrombus formation in the test system, which is associated with a reduction of the circulating blood cells.The reported in vitro dynamic whole blood test system allowed the sensitive analysis of the hemocompatibility of polymer-based microparticles and was successfully validated for porous PEI microparticles with different water wettabilities. Beyond the qualitative and quantitative analysis of cell-material interactions, the test also allowed the functional evaluation of platelets in whole blood.

Lung function, transfusion, pulmonary capillary blood volume and sickle cell disease.

PubMed

Lunt, Alan; McGhee, Emily; Robinson, Polly; Rees, David; Height, Susan; Greenough, Anne

2016-02-01

Lung function abnormalities occur in children with sickle cell disease (SCD) and may be associated with elevated pulmonary blood volume. To investigate that association, we determined whether blood transfusion in SCD children acutely increased pulmonary capillary blood volume (PCBV) and increased respiratory system resistance (Rrs5). Measurements of Rrs5 and spirometry were made before and after blood transfusion in 18 children, median age 14.2 (6.6-18.5) years. Diffusing capacity for carbon monoxide and nitric oxide were assessed to calculate the PCBV. Post transfusion, the median Rrs5 had increased from 127.4 to 141.3% predicted (p<0.0001) and pulmonary capillary blood volume from 39.7 to 64.1 ml/m2 (p<0.0001); forced expiratory volume in one second (p=0.0056) and vital capacity (p=0.0008) decreased. The increase in Rrs5 correlated with the increase in PCBV (r=0.50, p=0.0493). Increased pulmonary capillary blood volume may at least partially explain the lung function abnormalities in SCD children. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of infrared light on live blood cells: Role of β-carotene.

PubMed

Barkur, Surekha; Bankapur, Aseefhali; Chidangil, Santhosh; Mathur, Deepak

2017-06-01

We have utilized Raman tweezers to measure and assign micro-Raman spectra of optically trapped, live red blood cells (RBCs), white blood cells (WBCs) and platelets. Various types of WBCs- both granulocytes, lymphocytes, and their different types have been studied. The Raman bands are assigned to different biomolecules of blood cells. The Raman spectra thus obtained has been enabled detection of β-carotene in these blood cells, the spectral features of which act as a signature that facilitates experimental probing of the effect of 785nm laser light on different blood cells as a function of incident laser power in the mW range. The spectral changes that we obtain upon laser irradiation indicate that, both haemoglobin as well as the cell membrane sustains damage. In case of lymphocytes and platelets the peaks corresponding to β-carotene showed drastic changes. Thorough analysis of the spectral changes indicates possibility of free radical induced damage of β-carotene in lymphocytes and platelets. Among different blood cells, RBCs have a power threshold of only 10mW. The power threshold for other types of blood cells is somewhat higher, but always below about 30mW. These values are likely to serve as useful guides for Raman tweezers based experiments on live cells. Copyright © 2017. Published by Elsevier B.V.

Novel clinical uses for cord blood derived mesenchymal stromal cells.

PubMed

Olson, Amanda L; McNiece, Ian K

2015-06-01

Regenerative medicine offers new hope for many debilitating diseases that result in damage to tissues and organs. The concept is straightforward with replacement of damaged cells with new functional cells. However, most tissues and organs are complex structures involving multiple cell types, supportive structures, a microenvironment producing cytokines and growth factors and a vascular system to supply oxygen and other nutrients. Therefore repair, particularly in the setting of ischemic damage, may require delivery of multiple cell types providing new vessel formation, a new microenvironment and functional cells. The field of stem cell biology has identified a number of stem cell sources including embryonic stem cells and adult stem cells that offer the potential to replace virtually all functional cells of the body. The focus of this article is a discussion of the potential of mesenchymal stromal cells (MSCs) from cord blood (CB) for regenerative medicine approaches. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

White blood cell segmentation by circle detection using electromagnetism-like optimization.

PubMed

Cuevas, Erik; Oliva, Diego; Díaz, Margarita; Zaldivar, Daniel; Pérez-Cisneros, Marco; Pajares, Gonzalo

2013-01-01

Medical imaging is a relevant field of application of image processing algorithms. In particular, the analysis of white blood cell (WBC) images has engaged researchers from fields of medicine and computer vision alike. Since WBCs can be approximated by a quasicircular form, a circular detector algorithm may be successfully applied. This paper presents an algorithm for the automatic detection of white blood cells embedded into complicated and cluttered smear images that considers the complete process as a circle detection problem. The approach is based on a nature-inspired technique called the electromagnetism-like optimization (EMO) algorithm which is a heuristic method that follows electromagnetism principles for solving complex optimization problems. The proposed approach uses an objective function which measures the resemblance of a candidate circle to an actual WBC. Guided by the values of such objective function, the set of encoded candidate circles are evolved by using EMO, so that they can fit into the actual blood cells contained in the edge map of the image. Experimental results from blood cell images with a varying range of complexity are included to validate the efficiency of the proposed technique regarding detection, robustness, and stability.

White Blood Cell Segmentation by Circle Detection Using Electromagnetism-Like Optimization

PubMed Central

Oliva, Diego; Díaz, Margarita; Zaldivar, Daniel; Pérez-Cisneros, Marco; Pajares, Gonzalo

2013-01-01

Medical imaging is a relevant field of application of image processing algorithms. In particular, the analysis of white blood cell (WBC) images has engaged researchers from fields of medicine and computer vision alike. Since WBCs can be approximated by a quasicircular form, a circular detector algorithm may be successfully applied. This paper presents an algorithm for the automatic detection of white blood cells embedded into complicated and cluttered smear images that considers the complete process as a circle detection problem. The approach is based on a nature-inspired technique called the electromagnetism-like optimization (EMO) algorithm which is a heuristic method that follows electromagnetism principles for solving complex optimization problems. The proposed approach uses an objective function which measures the resemblance of a candidate circle to an actual WBC. Guided by the values of such objective function, the set of encoded candidate circles are evolved by using EMO, so that they can fit into the actual blood cells contained in the edge map of the image. Experimental results from blood cell images with a varying range of complexity are included to validate the efficiency of the proposed technique regarding detection, robustness, and stability. PMID:23476713

Distinct bone marrow blood vessels differentially regulate haematopoiesis.

PubMed

Itkin, Tomer; Gur-Cohen, Shiri; Spencer, Joel A; Schajnovitz, Amir; Ramasamy, Saravana K; Kusumbe, Anjali P; Ledergor, Guy; Jung, Yookyung; Milo, Idan; Poulos, Michael G; Kalinkovich, Alexander; Ludin, Aya; Kollet, Orit; Shakhar, Guy; Butler, Jason M; Rafii, Shahin; Adams, Ralf H; Scadden, David T; Lin, Charles P; Lapidot, Tsvee

2016-04-21

Bone marrow endothelial cells (BMECs) form a network of blood vessels that regulate both leukocyte trafficking and haematopoietic stem and progenitor cell (HSPC) maintenance. However, it is not clear how BMECs balance these dual roles, and whether these events occur at the same vascular site. We found that mammalian bone marrow stem cell maintenance and leukocyte trafficking are regulated by distinct blood vessel types with different permeability properties. Less permeable arterial blood vessels maintain haematopoietic stem cells in a low reactive oxygen species (ROS) state, whereas the more permeable sinusoids promote HSPC activation and are the exclusive site for immature and mature leukocyte trafficking to and from the bone marrow. A functional consequence of high permeability of blood vessels is that exposure to blood plasma increases bone marrow HSPC ROS levels, augmenting their migration and differentiation, while compromising their long-term repopulation and survival. These findings may have relevance for clinical haematopoietic stem cell transplantation and mobilization protocols.

Cryopreservation has no effect on function of natural killer cells differentiated in vitro from umbilical cord blood CD34(+) cells.

PubMed

Domogala, Anna; Madrigal, J Alejandro; Saudemont, Aurore

2016-06-01

Natural killer (NK) cells offer the potential for a powerful cellular immunotherapy because they can target malignant cells without being direct effectors of graft-versus-host disease. We have previously shown that high numbers of functional NK cells can be differentiated in vitro from umbilical cord blood (CB) CD34(+) cells. To develop a readily available, off-the-shelf cellular product, it is essential that NK cells differentiated in vitro can be frozen and thawed while maintaining the same phenotype and functions. We evaluated the phenotype and function of fresh and frozen NK cells differentiated in vitro. We also assessed whether the concentration of NK cells at the time of freezing had an impact on cell viability. We found that cell concentration of NK cells at the time of freezing did not have an impact on their viability and on cell recovery post-thaw. Moreover, freezing of differentiated NK cells in vitro did not affect their phenotype, cytotoxicity and degranulation capacity toward K562 cells, cytokine production and proliferation. We are therefore able to generate large numbers of functional NK cells from CB CD34(+) cells that maintain the same phenotype and function post-cryopreservation, which will allow for multiple infusions of a highly cytotoxic NK cell product. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Surfactant Functionalization Induces Robust, Differential Adhesion of Tumor Cells and Blood Cells to Charged Nanotube-Coated Biomaterials Under Flow

PubMed Central

Mitchell, Michael J.; Castellanos, Carlos A.; King, Michael R.

2015-01-01

The metastatic spread of cancer cells from the primary tumor to distant sites leads to a poor prognosis in cancers originating from multiple organs. Increasing evidence has linked selectin-based adhesion between circulating tumor cells (CTCs) and endothelial cells of the microvasculature to metastatic dissemination, in a manner similar to leukocyte adhesion during inflammation. Functionalized biomaterial surfaces hold promise as a diagnostic tool to separate CTCs and potentially treat metastasis, utilizing antibody and selectin-mediated interactions for cell capture under flow. However, capture at high purity levels is challenged by the fact that CTCs and leukocytes both possess selectin ligands. Here, a straightforward technique to functionalize and alter the charge of naturally occurring halloysite nanotubes using surfactants is reported to induce robust, differential adhesion of tumor cells and blood cells to nanotube-coated surfaces under flow. Negatively charged sodium dodecanoate-functionalized nanotubes simultaneously enhanced tumor cell capture while negating leukocyte adhesion, both in the presence and absence of adhesion proteins, and can be utilized to isolate circulating tumor cells regardless of biomarker expression. Conversely, diminishing nanotube charge via functionalization with decyltrimethylammonium bromide both abolished tumor cell capture while promoting leukocyte adhesion. PMID:25934290

A functional requirement for astroglia in promoting blood vessel development in the early postnatal brain.

PubMed

Ma, Shang; Kwon, Hyo Jun; Huang, Zhen

2012-01-01

Astroglia are a major cell type in the brain and play a key role in many aspects of brain development and function. In the adult brain, astrocytes are known to intimately ensheath blood vessels and actively coordinate local neural activity and blood flow. During development of the neural retina, blood vessel growth follows a meshwork of astrocytic processes. Several genes have also been implicated in retinal astrocytes for regulating vessel development. This suggests a role of astrocytes in promoting angiogenesis throughout the central nervous system. To determine the roles that astrocytes may play during brain angiogenesis, we employ genetic approaches to inhibit astrogliogenesis during perinatal corticogenesis and examine its effects on brain vessel development. We find that conditional deletion from glial progenitors of orc3, a gene required for DNA replication, dramatically reduces glial progenitor cell number in the subventricular zone and astrocytes in the early postnatal cerebral cortex. This, in turn, results in severe reductions in both the density and branching frequency of cortical blood vessels. Consistent with a delayed growth but not regression of vessels, we find neither significant net decreases in vessel density between different stages after normalizing for cortical expansion nor obvious apoptosis of endothelial cells in these mutants. Furthermore, concomitant with loss of astroglial interactions, we find increased endothelial cell proliferation, enlarged vessel luminal size as well as enhanced cytoskeletal gene expression in pericytes, which suggests compensatory changes in vascular cells. Lastly, we find that blood vessel morphology in mutant cortices recovers substantially at later stages, following astrogliosis. These results thus implicate a functional requirement for astroglia in promoting blood vessel growth during brain development.

A nanostructure of functional targeting epirubicin liposomes dually modified with aminophenyl glucose and cyclic pentapeptide used for brain glioblastoma treatment

PubMed Central

Zhang, Cheng-Xiang; Zhao, Wei-Yu; Liu, Lei; Ju, Rui-Jun; Mu, Li-Min; Zhao, Yao; Zeng, Fan; Xie, Hong-Jun; Yan, Yan; Lu, Wan-Liang

2015-01-01

The objectives of the present study were to develop functional targeting epirubicin liposomes for transferring drugs across the blood-brain barrier (BBB), treating glioblastoma, and disabling neovascularization. The studies were performed on glioblastoma cells in vitro and on glioblastoma-bearing mice. The results showed that the constructed liposomes had a high encapsulation efficiency for drugs (>95%), suitable particle size (109 nm), and less leakage in the blood component-containing system; were significantly able to be transported across the BBB; and exhibited efficacies in killing glioblastoma cells and in destroying glioblastoma neovasculature in vitro and in glioblastoma-bearing mice. The action mechanisms of functional targeting epirubicin liposomes correlated with the following features: the long circulation in the blood system, the ability to be transported across the BBB via glucose transporter-1, and the targeting effects on glioblastoma cells and on the endothelial cells of the glioblastoma neovasculature via the integrin β3 receptor. In conclusion, functional targeting epirubicin liposomes could be used as a potential therapy for treating brain glioblastoma and disabling neovascularization in brain glioblastomas. PMID:26418720

Erythrocyte deformation in ischemic acute tubular necrosis and amelioration by splenectomy in the dog.

PubMed

Mandal, A K; Taylor, C A; Bell, R D; Hillman, N M; Jarnot, M D; Cunningham, J D; Phillips, L G

1991-11-01

Bilateral renal artery occlusion (RAO) for 120 minutes in dogs results in acute tubular necrosis (ATN) and peritubular capillary (PTC) congestion with rapidly deteriorating renal function. We have shown that prior splenectomy minimizes RAO-induced renal functional and histopathologic changes. The purpose of this study was to examine whether this renal protection is due to prevention of red blood cell echinocyte formation and resultant renal PTC congestion. Echinocytes (burr cells) are poorly deformable, impart high viscosity to the blood, and may hinder reperfusion by increasing resistance to renal capillary blood flow. Splenectomized (SPLX) or sham-SPLX dogs were treated with bilateral RAO for 120 minutes. After RAO, renal function and renal blood flow were monitored, and peripheral blood red blood cells were examined at 1 hour and at 24-hour intervals for 96 hours. Renal biopsies were taken 1 hour after RAO and the kidneys removed 96 hours after RAO. The RBCs and renal tissues were studied using scanning electron microscopy. Renal function was assessed by endogenous creatinine clearance. Sham-SPLX animals showed a marked and sustained decrease in creatinine clearance, consistently elevated serum creatinine levels and fractional excretion of sodium, and diffuse ATN and PTC congestion with echinocytes. These animals had a peak in circulating echinocytes 1 hour after RAO (p less than 0.05), which showed an excellent negative correlation with creatinine clearance (r = -0.999; p less than 0.001). On the contrary, SPLX animals had essentially no change in serum creatinine or fractional excretion of sodium, minimal tubular changes, no PTC congestion, and no rise in circulating echinocytes during the 96-hour observation. In vitro treatment of the postischemic red blood cells from sham animals with adenosine-inosine or fresh postischemic plasma from the SPLX animals showed almost complete reversal to discocytes (normal red blood cells), whereas in vitro treatment of postischemic red blood cells from the SPLX animals with fresh postischemic plasma from the sham animals resulted in a marked echinocytic response. We conclude that 1) a marked echinocyte response in the immediate postischemic period is an important mechanism in initiating ischemic ATN, 2) an echinocyte inducing factor may reside in the plasma of spleen-intact animals, and 3) mitigation of ATN and PTC congestion by splenectomy is, at least in part, consequential to attenuated echinocytic response in the immediate postischemic period.

Predicting human blood viscosity in silico

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fedosov, Dmitry A.; Pan, Wenxiao; Caswell, Bruce

2011-07-05

Cellular suspensions such as blood are a part of living organisms and their rheological and flow characteristics determine and affect majority of vital functions. The rheological and flow properties of cell suspensions are determined by collective dynamics of cells, their structure or arrangement, cell properties and interactions. We study these relations for blood in silico using a mesoscopic particle-based method and two different models (multi-scale/low-dimensional) of red blood cells. The models yield accurate quantitative predictions of the dependence of blood viscosity on shear rate and hematocrit. We explicitly model cell aggregation interactions and demonstrate the formation of reversible rouleaux structuresmore » resulting in a tremendous increase of blood viscosity at low shear rates and yield stress, in agreement with experiments. The non-Newtonian behavior of such cell suspensions (e.g., shear thinning, yield stress) is analyzed and related to the suspension’s microstructure, deformation and dynamics of single cells. We provide the flrst quantitative estimates of normal stress differences and magnitude of aggregation forces in blood. Finally, the flexibility of the cell models allows them to be employed for quantitative analysis of a much wider class of complex fluids including cell, capsule, and vesicle suspensions.« less

Pericytes of the neurovascular unit: Key functions and signaling pathways

PubMed Central

Sweeney, Melanie D.; Ayyadurai, Shiva; Zlokovic, Berislav V.

2017-01-01

Pericytes are vascular mural cells embedded in the basement membrane of blood microvessels. They extend their processes along capillaries, pre-capillary arterioles, and post-capillary venules. The central nervous system (CNS) pericytes are uniquely positioned within the neurovascular unit between endothelial cells, astrocytes, and neurons. They integrate, coordinate, and process signals from their neighboring cells to generate diverse functional responses that are critical for CNS functions in health and disease including regulation of the blood-brain barrier permeability, angiogenesis, clearance of toxic metabolites, capillary hemodynamic responses, neuroinflammation, and stem cell activity. Here, we examine the key signaling pathways between pericytes and their neighboring endothelial cells, astrocytes, and neurons that control neurovascular functions. We also review the role of pericytes in different CNS disorders including rare monogenic diseases and complex neurological disorders such as Alzheimer's disease and brain tumors. Finally, we discuss directions for future studies. PMID:27227366

Cord blood-derived CD34+ hematopoietic cells with low mitochondrial mass are enriched in hematopoietic repopulating stem cell function.

PubMed

Romero-Moya, Damia; Bueno, Clara; Montes, Rosa; Navarro-Montero, Oscar; Iborra, Francisco J; López, Luis Carlos; Martin, Miguel; Menendez, Pablo

2013-07-01

The homeostasis of the hematopoietic stem/progenitor cell pool relies on a fine-tuned balance between self-renewal, differentiation and proliferation. Recent studies have proposed that mitochondria regulate these processes. Although recent work has contributed to understanding the role of mitochondria during stem cell differentiation, it remains unclear whether the mitochondrial content/function affects human hematopoietic stem versus progenitor function. We found that mitochondrial mass correlates strongly with mitochondrial membrane potential in CD34(+) hematopoietic stem/progenitor cells. We, therefore, sorted cord blood CD34(+) cells on the basis of their mitochondrial mass and analyzed the in vitro homeostasis and clonogenic potential as well as the in vivo repopulating potential of CD34(+) cells with high (CD34(+) Mito(High)) versus low (CD34(+) Mito(Low)) mitochondrial mass. The CD34(+) Mito(Low) fraction contained 6-fold more CD34(+)CD38(-) primitive cells and was enriched in hematopoietic stem cell function, as demonstrated by its significantly greater hematopoietic reconstitution potential in immuno-deficient mice. In contrast, the CD34(+) Mito(High) fraction was more enriched in hematopoietic progenitor function with higher in vitro clonogenic capacity. In vitro differentiation of CD34(+) Mito(Low) cells was significantly delayed as compared to that of CD34(+) Mito(High) cells. The eventual complete differentiation of CD34(+) Mito(Low) cells, which coincided with a robust expansion of the CD34(-) differentiated progeny, was accompanied by mitochondrial adaptation, as shown by significant increases in ATP production and expression of the mitochondrial genes ND1 and COX2. In conclusion, cord blood CD34(+) cells with low levels of mitochondrial mass are enriched in hematopoietic repopulating stem cell function whereas high levels of mitochondrial mass identify hematopoietic progenitors. A mitochondrial response underlies hematopoietic stem/progenitor cell differentiation and proliferation of lineage-committed CD34(-) cells.

Investigation of nanodiamonds interactions in canine blood

NASA Astrophysics Data System (ADS)

WÄ sowicz, Michał; Marek, Kulka; Cićkiewicz, Maciej; Cymerman, Magdalena

2017-02-01

The whole blood contains red cells, white cells, and platelets suspended in plasma. In the following study we investigated an impact of nanodiamond particles on blood elements over various periods of time.The material used in the study consisted of samples taken from ten healthy canines (Canis lupus f. domestica) of various age, different blood types and both sexes. The markings were conducted by adding to the blood unmodified diamonds (SND), modified O2 (SO2) suspended in 0,9% NaCl. The blood was put under an impact of two diamond concentrations: 20μl and 100μl. The amount of abnormal cells increased with time. The percentage of echinocytes as a result of interaction with nanodiamonds in various time periods for individual specimens was scarce. In the examined microscopic image a summary was made for 100 white blood cells. Following cells were included in said group: band neutrophils, segmented neutrophils, eosinophils, basophils, lymphocytes, monocytes, lymphocytes with granulates, stimulated lymphocytes, lymphocytes with vacuoles, metamielocytes and smudge cells. The impact of the three diamond types had no clinical importance on red blood cells. After the diamonds mixed with white blood cells, atypical cells came into being, in the range of agranulocytes in stimulated form or with granulates and/or vacuoles. It is supposed that as a result of longlasting exposure a stimulation and vacuolisation takes place, because of the function of the cells.

Virtual Global Transplant Laboratory Standard Operating Procedures for Blood Collection, PBMC Isolation, and Storage.

PubMed

Higdon, Lauren E; Lee, Karim; Tang, Qizhi; Maltzman, Jonathan S

2016-09-01

Research on human immune responses frequently involves the use of peripheral blood mononuclear cells (PBMC) immediately, or at significantly delayed timepoints, after collection. This requires PBMC isolation from whole blood and cryopreservation for some applications. It is important to standardize protocols for blood collection, PBMC isolation, cryopreservation, and thawing that maximize survival and functionality of PBMC at the time of analysis. This resource includes detailed protocols describing blood collection tubes, isolation of PBMC using a density gradient, cryopreservation of PBMC, and thawing of cells as well as preparation for functional assays. For each protocol, we include important considerations, such as timing, storage temperatures, and freezing rate. In addition, we provide alternatives so that researchers can make informed decisions in determining the optimal protocol for their application.

Functional Comparison of Induced Pluripotent Stem Cell- and Blood-Derived GPIIbIIIa Deficient Platelets

PubMed Central

Haas, Jessica; Sandrock-Lang, Kirstin; Gärtner, Florian; Jung, Christian Billy; Zieger, Barbara; Parrotta, Elvira; Kurnik, Karin; Sinnecker, Daniel; Wanner, Gerhard; Laugwitz, Karl-Ludwig; Massberg, Steffen; Moretti, Alessandra

2015-01-01

Human induced pluripotent stem cells (hiPSCs) represent a versatile tool to model genetic diseases and are a potential source for cell transfusion therapies. However, it remains elusive to which extent patient-specific hiPSC-derived cells functionally resemble their native counterparts. Here, we generated a hiPSC model of the primary platelet disease Glanzmann thrombasthenia (GT), characterized by dysfunction of the integrin receptor GPIIbIIIa, and compared side-by-side healthy and diseased hiPSC-derived platelets with peripheral blood platelets. Both GT-hiPSC-derived platelets and their peripheral blood equivalents showed absence of membrane expression of GPIIbIIIa, a reduction of PAC-1 binding, surface spreading and adherence to fibrinogen. We demonstrated that GT-hiPSC-derived platelets recapitulate molecular and functional aspects of the disease and show comparable behavior to their native counterparts encouraging the further use of hiPSC-based disease models as well as the transition towards a clinical application. PMID:25607928

The Blood–Brain Barrier

PubMed Central

Daneman, Richard; Prat, Alexandre

2015-01-01

Blood vessels are critical to deliver oxygen and nutrients to all of the tissues and organs throughout the body. The blood vessels that vascularize the central nervous system (CNS) possess unique properties, termed the blood–brain barrier, which allow these vessels to tightly regulate the movement of ions, molecules, and cells between the blood and the brain. This precise control of CNS homeostasis allows for proper neuronal function and also protects the neural tissue from toxins and pathogens, and alterations of these barrier properties are an important component of pathology and progression of different neurological diseases. The physiological barrier is coordinated by a series of physical, transport, and metabolic properties possessed by the endothelial cells (ECs) that form the walls of the blood vessels, and these properties are regulated by interactions with different vascular, immune, and neural cells. Understanding how these different cell populations interact to regulate the barrier properties is essential for understanding how the brain functions during health and disease. PMID:25561720

Modelling the endothelial blood-CNS barriers: a method for the production of robust in vitro models of the rat blood-brain barrier and blood-spinal cord barrier

PubMed Central

2013-01-01

Background Modelling the blood-CNS barriers of the brain and spinal cord in vitro continues to provide a considerable challenge for research studying the passage of large and small molecules in and out of the central nervous system, both within the context of basic biology and for pharmaceutical drug discovery. Although there has been considerable success over the previous two decades in establishing useful in vitro primary endothelial cell cultures from the blood-CNS barriers, no model fully mimics the high electrical resistance, low paracellular permeability and selective influx/efflux characteristics of the in vivo situation. Furthermore, such primary-derived cultures are typically labour-intensive and generate low yields of cells, limiting scope for experimental work. We thus aimed to establish protocols for the high yield isolation and culture of endothelial cells from both rat brain and spinal cord. Our aim was to optimise in vitro conditions for inducing phenotypic characteristics in these cells that were reminiscent of the in vivo situation, such that they developed into tight endothelial barriers suitable for performing investigative biology and permeability studies. Methods Brain and spinal cord tissue was taken from the same rats and used to specifically isolate endothelial cells to reconstitute as in vitro blood-CNS barrier models. Isolated endothelial cells were cultured to expand the cellular yield and then passaged onto cell culture inserts for further investigation. Cell culture conditions were optimised using commercially available reagents and the resulting barrier-forming endothelial monolayers were characterised by functional permeability experiments and in vitro phenotyping by immunocytochemistry and western blotting. Results Using a combination of modified handling techniques and cell culture conditions, we have established and optimised a protocol for the in vitro culture of brain and, for the first time in rat, spinal cord endothelial cells. High yields of both CNS endothelial cell types can be obtained, and these can be passaged onto large numbers of cell culture inserts for in vitro permeability studies. The passaged brain and spinal cord endothelial cells are pure and express endothelial markers, tight junction proteins and intracellular transport machinery. Further, both models exhibit tight, functional barrier characteristics that are discriminating against large and small molecules in permeability assays and show functional expression of the pharmaceutically important P-gp efflux transporter. Conclusions Our techniques allow the provision of high yields of robust sister cultures of endothelial cells that accurately model the blood-CNS barriers in vitro. These models are ideally suited for use in studying the biology of the blood-brain barrier and blood-spinal cord barrier in vitro and for pre-clinical drug discovery. PMID:23773766

Modelling the endothelial blood-CNS barriers: a method for the production of robust in vitro models of the rat blood-brain barrier and blood-spinal cord barrier.

PubMed

Watson, P Marc D; Paterson, Judy C; Thom, George; Ginman, Ulrika; Lundquist, Stefan; Webster, Carl I

2013-06-18

Modelling the blood-CNS barriers of the brain and spinal cord in vitro continues to provide a considerable challenge for research studying the passage of large and small molecules in and out of the central nervous system, both within the context of basic biology and for pharmaceutical drug discovery. Although there has been considerable success over the previous two decades in establishing useful in vitro primary endothelial cell cultures from the blood-CNS barriers, no model fully mimics the high electrical resistance, low paracellular permeability and selective influx/efflux characteristics of the in vivo situation. Furthermore, such primary-derived cultures are typically labour-intensive and generate low yields of cells, limiting scope for experimental work. We thus aimed to establish protocols for the high yield isolation and culture of endothelial cells from both rat brain and spinal cord. Our aim was to optimise in vitro conditions for inducing phenotypic characteristics in these cells that were reminiscent of the in vivo situation, such that they developed into tight endothelial barriers suitable for performing investigative biology and permeability studies. Brain and spinal cord tissue was taken from the same rats and used to specifically isolate endothelial cells to reconstitute as in vitro blood-CNS barrier models. Isolated endothelial cells were cultured to expand the cellular yield and then passaged onto cell culture inserts for further investigation. Cell culture conditions were optimised using commercially available reagents and the resulting barrier-forming endothelial monolayers were characterised by functional permeability experiments and in vitro phenotyping by immunocytochemistry and western blotting. Using a combination of modified handling techniques and cell culture conditions, we have established and optimised a protocol for the in vitro culture of brain and, for the first time in rat, spinal cord endothelial cells. High yields of both CNS endothelial cell types can be obtained, and these can be passaged onto large numbers of cell culture inserts for in vitro permeability studies. The passaged brain and spinal cord endothelial cells are pure and express endothelial markers, tight junction proteins and intracellular transport machinery. Further, both models exhibit tight, functional barrier characteristics that are discriminating against large and small molecules in permeability assays and show functional expression of the pharmaceutically important P-gp efflux transporter. Our techniques allow the provision of high yields of robust sister cultures of endothelial cells that accurately model the blood-CNS barriers in vitro. These models are ideally suited for use in studying the biology of the blood-brain barrier and blood-spinal cord barrier in vitro and for pre-clinical drug discovery.

An antibody to the lutheran glycoprotein (Lu) recognizing the LU4 blood type variant inhibits cell adhesion to laminin α5.

PubMed

Kikkawa, Yamato; Miwa, Takahiro; Tohara, Yukiko; Hamakubo, Takayuki; Nomizu, Motoyoshi

2011-01-01

The Lutheran blood group glycoprotein (Lu), an Ig superfamily (IgSF) transmembrane receptor, is also known as basal cell adhesion molecule (B-CAM). Lu/B-CAM is a specific receptor for laminin α5, a major component of basement membranes in various tissues. Previous reports have shown that Lu/B-CAM binding to laminin α5 contributes to sickle cell vaso-occlusion. However, as there are no useful tools such as function-blocking antibodies or drugs, it is unclear how epithelial and sickled red blood cells adhere to laminin α5 via Lu/B-CAM. In this study, we discovered a function-blocking antibody that inhibits Lu binding to laminin α5 using a unique binding assay on tissue sections. To characterize the function-blocking antibody, we identified the site on Lu/B-CAM recognized by this antibody. The extracellular domain of Lu/B-CAM contains five IgSF domains, D1-D2-D3-D4-D5. The antibody epitope was localized to D2, but not to the D3 domain containing the major part of the laminin α5 binding site. Furthermore, mutagenesis studies showed that Arg(175), the LU4 blood group antigenic site, was crucial for forming the epitope and the antibody bound sufficiently close to sterically hinder the interaction with α5. Cell adhesion assay using the antibody also showed that Lu/B-CAM serves as a secondary receptor for the adhesion of carcinoma cells to laminin α5. This function-blocking antibody against Lu/B-CAM should be useful for not only investigating cell adhesion to laminin α5 but also for developing drugs to inhibit sickle cell vaso-occlusion.

Surface-Micromachined Microfiltration Membranes for Efficient Isolation and Functional Immunophenotyping of Subpopulations of Immune Cells

PubMed Central

Oh, Boram; Lam, Raymond H. W.; Fan, Rong; Cornell, Timothy T.; Shanley, Thomas P.; Kurabayashi, Katsuo; Fu, Jianping

2015-01-01

An accurate measurement of the immune status in patients with immune system disorders is critical in evaluating the stage of diseases and tailoring drug treatments. The functional cellular immunity test is a promising method to establish the diagnosis of immune dysfunctions. The conventional functional cellular immunity test involves measurements of the capacity of peripheral blood mononuclear cells to produce pro-inflammatory cytokines when stimulated ex vivo. However, this “bulk” assay measures the overall reactivity of a population of lymphocytes and monocytes, making it difficult to pinpoint the phenotype or real identity of the reactive immune cells involved. In this research, we develop a large surface micromachined polydimethylsiloxane (PDMS) microfiltration membrane (PMM) with high porosity, which is integrated in a microfluidic microfiltration platform. Using the PMM with functionalized microbeads conjugated with antibodies against specific cell surface proteins, we demonstrated rapid, efficient and high-throughput on-chip isolation, enrichment, and stimulation of subpopulations of immune cells from blood specimens. Furthermore, the PMM-integrated microfiltration platform, coupled with a no-wash homogeneous chemiluminescence assay (“AlphaLISA”), enables us to demonstrate rapid and sensitive on-chip immunophenotyping assays for subpopulations of immune cells isolated directly from minute quantities of blood samples. PMID:23335389

Novel single-cell functional analysis of red blood cells using laser tweezers Raman spectroscopy: application for sickle cell disease.

PubMed

Liu, Rui; Mao, Ziliang; Matthews, Dennis L; Li, Chin-Shang; Chan, James W; Satake, Noriko

2013-07-01

Laser tweezers Raman spectroscopy was used to characterize the oxygenation response of single normal adult, sickle, and cord blood red blood cells (RBCs) to an applied mechanical force. Individual cells were subjected to different forces by varying the laser power of a single-beam optical trap, and the intensities of several oxygenation-specific Raman spectral peaks were monitored to determine the oxygenation state of the cells. For all three cell types, an increase in laser power (or mechanical force) induced a greater deoxygenation of the cell. However, sickle RBCs deoxygenated more readily than normal RBCs when subjected to the same optical forces. Conversely, cord blood RBCs were able to maintain their oxygenation better than normal RBCs. These results suggest that differences in the chemical or mechanical properties of fetal, normal, and sickle cells affect the degree to which applied mechanical forces can deoxygenate the cell. Populations of normal, sickle, and cord RBCs were identified and discriminated based on this mechanochemical phenomenon. This study demonstrates the potential application of laser tweezers Raman spectroscopy as a single-cell, label-free analytical tool to characterize the functional (e.g., mechanical deformability, oxygen binding) properties of normal and diseased RBCs. Copyright © 2013 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.

Early diagnosis of diabetic vascular complications: impairment of red blood cell deformability

NASA Astrophysics Data System (ADS)

Shin, Sehyun; Ku, Yunhee; Park, Cheol-Woo; Suh, Jang-Soo

2006-02-01

Reduced deformability of red blood cells (RBCs) may play an important role on the pathogenesis of chronic vascular complications of diabetes mellitus. However, available techniques for measuring RBC deformability often require washing process after each measurement, which is not optimal for day-to-day clinical use at point of care. The objectives of the present study are to develop a device and to delineate the correlation of impaired RBC deformability with diabetic nephropathy. We developed a disposable ektacytometry to measure RBC deformability, which adopted a laser diffraction technique and slit rheometry. The essential features of this design are its simplicity (ease of operation and no moving parts) and a disposable element which is in contact with the blood sample. We studied adult diabetic patients divided into three groups according to diabetic complications. Group I comprised 57 diabetic patients with normal renal function. Group II comprised 26 diabetic patients with chronic renal failure (CRF). Group III consisted of 30 diabetic subjects with end-stage renal disease (ESRD) on hemodialysis. According to the renal function for the diabetic groups, matched non-diabetic groups were served as control. We found substantially impaired red blood cell deformability in those with normal renal function (group I) compared to non-diabetic control (P = 0.0005). As renal function decreases, an increased impairment in RBC deformability was found. Diabetic patients with chronic renal failure (group II) when compared to non-diabetic controls (CRF) had an apparently greater impairment in RBC deformability (P = 0.07). The non-diabetic cohort (CRF), on the other hand, manifested significant impairment in red blood cell deformability compared to healthy control (P = 0.0001). The newly developed slit ektacytometer can measure the RBC deformability with ease and accuracy. In addition, progressive impairment in cell deformability is associated with renal function loss in all patients regardless of the presence or absence of diabetes. In diabetic patients, early impairment in RBC deformability appears in patients with normal renal function.

Generation of erythroid cells from polyploid giant cancer cells: re-thinking about tumor blood supply.

PubMed

Yang, Zhigang; Yao, Hong; Fei, Fei; Li, Yuwei; Qu, Jie; Li, Chunyuan; Zhang, Shiwu

2018-04-01

During development and tumor progression, cells need a sufficient blood supply to maintain development and rapid growth. It is reported that there are three patterns of blood supply for tumor growth: endothelium-dependent vessels, mosaic vessels, and vasculogenic mimicry (VM). VM was first reported in highly aggressive uveal melanomas, with tumor cells mimicking the presence and function of endothelial cells forming the walls of VM vessels. The walls of mosaic vessels are randomly lined with both endothelial cells and tumor cells. We previously proposed a three-stage process, beginning with VM, progressing to mosaic vessels, and eventually leading to endothelium-dependent vessels. However, many phenomena unique to VM channel formation remain to be elucidated, such as the origin of erythrocytes before VM vessels connect with endothelium-dependent vessels. In adults, erythroid cells are generally believed to be generated from hematopoietic stem cells in the bone marrow. In contrast, embryonic tissue obtains oxygen through formation of blood islands, which are largely composed of embryonic hemoglobin with a higher affinity with oxygen, in the absence of mature erythrocytes. Recent data from our laboratory suggest that embryonic blood-forming mechanisms also exist in cancer tissue, particularly when these tissues are under environmental stress such as hypoxia. We review the evidence from induced pluripotent stem cells in vitro and in vivo to support this previously underappreciated cell functionality in normal and cancer cells, including the ability to generate erythroid cells. We will also summarize the current understanding of tumor angiogenesis, VM, and our recent work on polyploid giant cancer cells, with emphasis on their ability to generate erythroid cells and their association with tumor growth under hypoxia. An alternative embryonic pathway to obtain oxygen in cancer cells exists, particularly when they are under hypoxic conditions.

Antigenic and functional properties of the human red blood cell urea transporter hUT-B1.

PubMed

Lucien, Nicole; Sidoux-Walter, Frédéric; Roudier, Nathalie; Ripoche, Pierre; Huet, Martine; Trinh-Trang-Tan, Marie-Marcelle; Cartron, Jean-Pierre; Bailly, Pascal

2002-09-13

The Kidd (JK) blood group locus encodes the urea transporter hUT-B1, which is expressed on human red blood cells and other tissues. The common JK*A/JK*B blood group polymorphism is caused by a single nucleotide transition G838A changing Asp-280 to Asn-280 on the polypeptide, and transfection of erythroleukemic K562 cells with hUT-B1 cDNAs carrying either the G838 or the A838 nucleotide substitutions resulted in the isolation of stable clones that expressed the Jk(a) or Jk(b) antigens, respectively, thus providing the first direct demonstration that the hUT-B1 gene encodes the Kidd blood group antigens. In addition, immunochemical analysis of red blood cells demonstrated that hUT-B1 also exhibits ABO determinants attached to the single N-linked sugar chain at Asn-211. Moreover, immunoadsorption studies, using inside-out and right-side-out red cell membrane vesicles as competing antigen, demonstrated that the C- and N-terminal ends of hUT-B1 are oriented intracellularly. Mutagenesis and functional studies by expression in Xenopus oocytes revealed that both cysteines Cys-25 and Cys-30 (but not alone) are essential for plasma membrane addressing. Conversely, the transport function was not affected by the JK*A/JK*B polymorphism, C-terminal deletion (residues 360-389), or mutation of the extracellular N-glycosylation consensus site and remains poorly para-chloromercuribenzene sulfonate (pCMBS)-sensitive. However, transport studies by stopped flow light scattering using Jk-K562 transfectants demonstrated that the hUT-B1-mediated urea transport is pCMBS-sensitive in an erythroid context, as reported previously for the transporter of human red blood cells. Mutagenesis analysis also indicated that Cys-151 and Cys-236, at least alone, are not involved in pCMBS inhibition. Altogether, these antigenic, topologic, and functional properties might have implications into the physiology of hUT-B1 and other members of the urea transporter family.

Dual role of wingless signaling in stem-like hematopoietic precursor maintenance in Drosophila.

PubMed

Sinenko, Sergey A; Mandal, Lolitika; Martinez-Agosto, Julian A; Banerjee, Utpal

2009-05-01

In Drosophila, blood development occurs in a specialized larval hematopoietic organ, the lymph gland (LG), within which stem-like hemocyte precursors or prohemocytes differentiate to multiple blood cell types. Here we show that components of the Wingless (Wg) signaling pathway are expressed in prohemocytes. Loss- and gain-of-function analysis indicates that canonical Wg signaling is required for maintenance of prohemocytes and negatively regulates their differentiation. Wg signals locally in a short-range fashion within different compartments of the LG. In addition, Wg signaling positively regulates the proliferation and maintenance of cells that function as a hematopoietic niche in Drosophila, the posterior signaling center (PSC), and in the proliferation of crystal cells. Our studies reveal a conserved function of Wg signaling in the maintenance of stem-like blood progenitors and reveal an involvement of this pathway in the regulation of hemocyte differentiation through its action in the hematopoietic niche.

Generation of dendritic cells from human bone marrow mononuclear cells: advantages for clinical application in comparison to peripheral blood monocyte derived cells.

PubMed

Bai, L; Feuerer, M; Beckhove, P; Umansky, V; Schirrmacher, V

2002-02-01

Dendritic cells (DCs) currently used for vaccination in clinical studies to induce immunity against malignant cells are normally generated from peripheral blood-derived monocytes. Here we studied conditions for the generation of DCs from unseparated human bone marrow (BM) mononuclear cells and compared them functionally with DCs from blood. The two types of DCs, from bone marrow (BM-DC) and peripheral blood (BL-DC), were generated in parallel from the same normal healthy donors by culturing in serum-free X-VIVO 20 medium containing GM-CSF and IL-4, and then the phenotypes and functions were compared. BM-DC generation occurred in 14 days and involved proliferative expansion from CD34 stem cells and differentiation while BL-DC generation occurred in 7 days from CD14 monocytes and involved only differentiation. A 7- to 25-fold higher number of DCs could be obtained from BM than from blood. BM-DC had similar phenotypes as BL-DC. The capacity to stimulate MLR reactivity in allogeneic T lymphocytes was higher with BM-DC than that with BL-DC. Also, the capacity to stimulate autologous memory T cell responses to tetanus toxoid (TT) or tuberculin (PPD) was higher with BM-DC than with BL-DC. These results suggest that BM-DC as produced here may be a very economic and useful source of professional antigen-presenting cells for anti-tumor immunotherapeutic protocols.

Measurement of Blood Coagulation Factor Synthesis in Cultures of Human Hepatocytes.

PubMed

Heinz, Stefan; Braspenning, Joris

2015-01-01

An important function of the liver is the synthesis and secretion of blood coagulation factors. Within the liver, hepatocytes are involved in the synthesis of most blood coagulation factors, such as fibrinogen, prothrombin, factor V, VII, IX, X, XI, XII, as well as protein C and S, and antithrombin, whereas liver sinusoidal endothelial cells produce factor VIII and von Willebrand factor. Here, we describe methods for the detection and quantification of most blood coagulation factors in hepatocytes in vitro. Hepatocyte cultures indeed provide a valuable tool to study blood coagulation factors. In addition, the generation and expansion of hepatocytes or hepatocyte-like cells may be used in future for cell-based therapies of liver diseases, including blood coagulation factor deficiencies.

Pretransplantation recipient regulatory T cell suppressive function predicts delayed and slow graft function after kidney transplantation.

PubMed

Nguyen, Minh-Tri J P; Fryml, Elise; Sahakian, Sossy K; Liu, Shuqing; Michel, Rene P; Lipman, Mark L; Mucsi, Istvan; Cantarovich, Marcelo; Tchervenkov, Jean I; Paraskevas, Steven

2014-10-15

Delayed graft function (DGF) and slow graft function (SGF) are a continuous spectrum of ischemia-reperfusion-related acute kidney injury (AKI) that increases the risk for acute rejection and graft loss after kidney transplantation. Regulatory T cells (Tregs) are critical in transplant tolerance and attenuate murine AKI. In this prospective observational cohort study, we evaluated whether pretransplantation peripheral blood recipient Treg frequency and suppressive function are predictors of DGF and SGF after kidney transplantation. Deceased donor kidney transplant recipients (n=53) were divided into AKI (n=37; DGF, n=10; SGF, n=27) and immediate graft function (n=16) groups. Pretransplantation peripheral blood CD4CD25FoxP3 Treg frequency was quantified by flow cytometry. Regulatory T-cell suppressive function was measured by suppression of autologous effector T-cell proliferation by Treg in co-culture. Pretransplantation Treg suppressive function, but not frequency, was decreased in AKI recipients (P<0.01). In univariate and multivariate analyses accounting for the effects of cold ischemic time and donor age, Treg suppressive function discriminated DGF from immediate graft function recipients in multinomial logistic regression (odds ratio, 0.77; P<0.01), accurately predicted AKI in receiver operating characteristic curve (area under the curve, 0.82; P<0.01), and predicted 14-day estimated glomerular filtration rate in linear regression (P<0.01). Our results indicate that recipient peripheral blood Treg suppressive function is a potential independent pretransplantation predictor of DGF and SGF.

Connexin 36 mediates blood cell flow in mouse pancreatic islets

PubMed Central

Short, Kurt W.; Head, W. Steve

2013-01-01

The insulin-secreting β-cells are contained within islets of Langerhans, which are highly vascularized. Blood cell flow rates through islets are glucose-dependent, even though there are no changes in blood cell flow within in the surrounding exocrine pancreas. This suggests a specific mechanism of glucose-regulated blood flow in the islet. Pancreatic islets respond to elevated glucose with synchronous pulses of electrical activity and insulin secretion across all β-cells in the islet. Connexin 36 (Cx36) gap junctions between islet β-cells mediate this synchronization, which is lost in Cx36 knockout mice (Cx36−/−). This leads to glucose intolerance in these mice, despite normal plasma insulin levels and insulin sensitivity. Thus, we sought to investigate whether the glucose-dependent changes in intraislet blood cell flow are also dependent on coordinated pulsatile electrical activity. We visualized and quantified blood cell flow using high-speed in vivo fluorescence imaging of labeled red blood cells and plasma. With the use of a live animal glucose clamp, blood cell flow was measured during either hypoglycemia (∼50 mg/dl) or hyperglycemia (∼300 mg/dl). In contrast to the large glucose-dependent islet blood velocity changes observed in wild-type mice, only minimal differences are observed in both Cx36+/− and Cx36−/− mice. This observation supports a novel model where intraislet blood cell flow is regulated by the coordinated electrical activity in the islet β-cells. Because Cx36 expression and function is reduced in type 2 diabetes, the resulting defect in intraislet blood cell flow regulation may also play a significant role in diabetic pathology. PMID:24326425

Extraction of immune and inflammatory cells from human lung parenchyma: evaluation of an enzymatic digestion procedure.

PubMed Central

Holt, P G; Robinson, B W; Reid, M; Kees, U R; Warton, A; Dawson, V H; Rose, A; Schon-Hegrad, M; Papadimitriou, J M

1986-01-01

The inflammatory and immune cell populations of the human lung parenchyma have not been characterized in detail. This report describes a novel and efficient procedure for their extraction. Histologically normal human lung tissue samples from pneumonectomy specimens were sliced to 0.5 mm, and digested in collagenase/DNAse. Viable mononuclear cell yields ranged from 15-48 X 10(6)/g, and were markedly in excess of reported methods employing mechanical tissue disruption, which normally yield populations containing almost exclusively macrophages. The lung digest population was examined by flow cytometry using monoclonal antibodies against cell surface receptors, and found to comprise up to 40% T lymphocytes, 10% B lymphocytes and 30% macrophages, contaminated by less than 1% peripheral blood cells. Based upon these figures, the recoverable lung parenchymal lymphoid cell pool appears considerably larger than previously recognized, being of the same order as the peripheral blood pool. Initial functional studies suggest that such cellular activities as antigen-specific T cell proliferation, antigen-presentation, interleukin 1 production and natural killer cell activity survive the extraction process, and controlled enzymatic digestion experiments with peripheral blood cells indicate that the degree of enzyme-mediated damage to these functions and to cell-surface structures, was minimal. The extraction method thus appears suitable for studying the types and functions of human parenchymal lung cells in health and disease. Images Fig. 2 p195-a PMID:3026698

Glucose-dependent blood flow dynamics in murine pancreatic islets in vivo

PubMed Central

Nyman, Lara R.; Ford, Eric

2010-01-01

Pancreatic islets are highly vascularized and arranged so that regions containing β-cells are distinct from those containing other cell types. Although islet blood flow has been studied extensively, little is known about the dynamics of islet blood flow during hypoglycemia or hyperglycemia. To investigate changes in islet blood flow as a function of blood glucose level, we clamped blood glucose sequentially at hyperglycemic (∼300 mg/dl or 16.8 mM) and hypoglycemic (∼50 mg/dl or 2.8 mM) levels while simultaneously imaging intraislet blood flow in mouse models that express green fluorescent protein in the β-cells or yellow fluorescent protein in the α-cells. Using line scanning confocal microscopy, in vivo blood flow was assayed after intravenous injection of fluorescent dextran or sulforhodamine-labeled red blood cells. Regardless of the sequence of hypoglycemia and hyperglycemia, islet blood flow is faster during hyperglycemia, and apparent blood volume is greater during hyperglycemia than during hypoglycemia. However, there is no change in the order of perfusion of different islet endocrine cell types in hypoglycemia compared with hyperglycemia, with the islet core of β-cells usually perfused first. In contrast to the results in islets, there was no significant difference in flow rate in the exocrine pancreas during hyperglycemia compared with hypoglycemia. These results indicate that glucose differentially regulates blood flow in the pancreatic islet vasculature independently of blood flow in the rest of the pancreas. PMID:20071562

Long-term Effects on the Histology and Function of Livers and Spleens in Rats after 33% Toploading of PEG-PLA-nano Artificial Red Blood Cells

PubMed Central

Liu, Zun Chang; Chang, Thomas M.S.

2012-01-01

This study is to investigate the long-term effects of nanodimension PEG-PLA artificial red blood cells containing hemoglobin and red blood cell enzymes on the liver and spleen after 1/3 blood volume top loading in rats. The experimental rats received one of the following infusions: Nano artificial red blood cells in Ringer lactate, Ringer lactate, stroma-free hemoglobin, polyhemoglobin, and autologous rat whole blood. Blood samples were taken before infusions and on days 1, 7, and 21 after infusions for analysis. Nano artificial red blood cells, polyhemoglobin, Ringer lactate and rat red blood cells did not have any significant adverse effects on alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, creatine kinase, amylase and creatine kinase. On the other hand, stroma-free hemoglobin induced significant adverse effects on liver as shown by elevation in alanine aminotransferase and aspartate aminotransferase throughout the 21 days. On day 21 after infusions rats were sacrificed and livers and spleens were excised for histological examination. Nano artificial red blood cells, polyhemoglobin, Ringer lactate and rat red blood cells did not cause any abnormalities in the microscopic histology of the livers and spleens. In the stroma-free hemoglobin group the livers showed accumulation of hemoglobin in central veins and sinusoids, and hepatic steatosis. In conclusion, injected nano artificial red blood cells can be efficiently metabolized and removed by the reticuloendothelial system, and do not have any biochemical or histological adverse effects on the livers or the spleens. PMID:19043818

Angiocrine functions of organ-specific endothelial cells

PubMed Central

Rafii, Shahin; Butler, Jason M; Ding, Bi-Sen

2016-01-01

Preface Endothelial cells lining blood vessel capillaries are not just passive conduits for delivering blood. Tissue-specific endothelium establish specialized vascular niches that deploy specific sets of growth factors, known as angiocrine factors, which actively participate in inducing, specifying, patterning, and guiding organ regeneration and maintaining homeostasis and metabolism. Angiocrine factors upregulated in response to injury orchestrates self-renewal and differentiation of tissue-specific repopulating resident stem and progenitor cells into functional organs. Uncovering the precise mechanisms whereby physiological-levels of angiocrine factors are spatially and temporally produced, and distributed by organotypic endothelium to repopulating cells, will lay the foundation for driving organ repair without scarring. PMID:26791722

Development of a human adaptive immune system in cord blood cell-transplanted mice.

PubMed

Traggiai, Elisabetta; Chicha, Laurie; Mazzucchelli, Luca; Bronz, Lucio; Piffaretti, Jean-Claude; Lanzavecchia, Antonio; Manz, Markus G

2004-04-02

Because ethical restrictions limit in vivo studies of the human hemato-lymphoid system, substitute human to small animal xenotransplantation models have been employed. Existing models, however, sustain only limited development and maintenance of human lymphoid cells and rarely produce immune responses. Here we show that intrahepatic injection of CD34+ human cord blood cells into conditioned newborn Rag2-/-gammac-/- mice leads to de novo development of B, T, and dendritic cells; formation of structured primary and secondary lymphoid organs; and production of functional immune responses. This provides a valuable model to study development and function of the human adaptive immune system in vivo.

Systemic treatment of focal brain injury in the rat by human umbilical cord blood cells being at different level of neural commitment.

PubMed

Gornicka-Pawlak, El Bieta; Janowski, Miroslaw; Habich, Aleksandra; Jablonska, Anna; Drela, Katarzyna; Kozlowska, Hanna; Lukomska, Barbara; Sypecka, Joanna; Domanska-Janik, Krystyna

2011-01-01

The aim of the study was to evaluate therapeutic effectiveness of intra-arterial infusion of human umbilical cord blood (HUCB) derived cells at different stages of their neural conversion. Freshly isolated mononuclear cells (D-0), neurally directed progenitors (D-3) and neural-like stem cells derived from umbilical cord blood (NSC) were compared. Focal brain damage was induced in rats by stereotactic injection of ouabain into dorsolateral striatum Three days later 10(7) of different subsets of HUCB cells were infused into the right internal carotid artery. Following surgery rats were housed in enriched environment for 30 days. Behavioral assessment consisted of tests for sensorimotor deficits (walking beam, rotarod, vibrissae elicited forelimb placing, apomorphine induced rotations), cognitive impairments (habit learning and object recognition) and exploratory behavior (open field). Thirty days after surgery the lesion volume was measured and the presence of donor cells was detected in the brain at mRNA level. At the same time immunohistochemical analysis of brain tissue was performed to estimate the local tissue response of ouabain injured rats and its modulation after HUCB cells systemic treatment. Functional effects of different subsets of cord blood cells shared substantial diversity in various behavioral tests. An additional analysis showed that D-0 HUCB cells were the most effective in functional restoration and reduction of brain lesion volume. None of transplanted cord blood derived cell fractions were detected in rat's brains at 30(th) day after treatment. This may suggest that the mechanism(s) underlying positive effects of HUCB derived cell may concern the other than direct neural cell supplementation. In addition increased immunoreactivity of markers indicating local cells proliferation and migration suggests stimulation of endogenous reparative processes by HUCB D-0 cell interarterial infusion.

Leukemic Cells "Gas Up" Leaky Bone Marrow Blood Vessels.

PubMed

Itkin, Tomer; Rafii, Shahin

2017-09-11

In this issue of Cancer Cell, Passaro et al. demonstrate how leukemia through aberrant induction of reactive oxygen species and nitric oxide production trigger marrow vessel leakiness, instigating pro-leukemic function. Disrupted tumor blood vessels promote exhaustion of non-malignant stem and progenitor cells and may facilitate leukemia relapse following chemotherapeutic treatment. Copyright © 2017. Published by Elsevier Inc.

[Correlations between functional activity of animal blood lymphocytes and change in solar activity].

PubMed

Karnaukhova, N A; Sergievich, L A

1999-01-01

It is shown that increase of Solar activity as measurement of the intensity of solar radio emissions at frequency of 2804 MHz leads to the reducing of the functional activity of immunocompetent cells in animal blood defining by parameter alpha.

A comparison of blood nitric oxide metabolites and hemoglobin functional properties among diving mammals.

PubMed

Fago, Angela; Parraga, Daniel Garcia; Petersen, Elin E; Kristensen, Niels; Giouri, Lea; Jensen, Frank B

2017-03-01

The ability of marine mammals to hunt prey at depth is known to rely on enhanced oxygen stores and on selective distribution of blood flow, but the molecular mechanisms regulating blood flow and oxygen transport remain unresolved. To investigate the molecular mechanisms that may be important in regulating blood flow, we measured concentration of nitrite and S-nitrosothiols (SNO), two metabolites of the vasodilator nitric oxide (NO), in the blood of 5 species of marine mammals differing in their dive duration: bottlenose dolphin, South American sea lion, harbor seal, walrus and beluga whale. We also examined oxygen affinity, sensitivity to 2,3-diphosphoglycerate (DPG) and nitrite reductase activity of the hemoglobin (Hb) to search for possible adaptive variations in these functional properties. We found levels of plasma and red blood cells nitrite similar to those reported for terrestrial mammals, but unusually high concentrations of red blood cell SNO in bottlenose dolphin, walrus and beluga whale, suggesting enhanced SNO-dependent signaling in these species. Purified Hbs showed similar functional properties in terms of oxygen affinity and sensitivity to DPG, indicating that reported large variations in blood oxygen affinity among diving mammals likely derive from phenotypic variations in red blood cell DPG levels. The nitrite reductase activities of the Hbs were overall slightly higher than that of human Hb, with the Hb of beluga whale, capable of longest dives, having the highest activity. Taken together, these results underscore adaptive variations in circulatory NO metabolism in diving mammals but not in the oxygenation properties of the Hb. Copyright © 2016 Elsevier Inc. All rights reserved.

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection

PubMed Central

Gordon, Claire L.; Thome, Joseph J.C.; Igarashi, Suzu

2017-01-01

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. PMID:28130404

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection.

PubMed

Gordon, Claire L; Miron, Michelle; Thome, Joseph J C; Matsuoka, Nobuhide; Weiner, Joshua; Rak, Michael A; Igarashi, Suzu; Granot, Tomer; Lerner, Harvey; Goodrum, Felicia; Farber, Donna L

2017-03-06

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. @Gordon et al.

Advances in small animal mesentery models for in vivo flow cytometry, dynamic microscopy, and drug screening

PubMed Central

Galanzha, Ekaterina I; Tuchin, Valery V; Zharov, Vladimir P

2007-01-01

Using animal mesentery with intravital optical microscopy is a well-established experimental model for studying blood and lymph microcirculation in vivo. Recent advances in cell biology and optical techniques provide the basis for extending this model for new applications, which should generate significantly improved experimental data. This review summarizes the achievements in this specific area, including in vivo label-free blood and lymph photothermal flow cytometry, super-sensitive fluorescence image cytometry, light scattering and speckle flow cytometry, microvessel dynamic microscopy, infrared (IR) angiography, and high-speed imaging of individual cells in fast flow. The capabilities of these techniques, using the rat mesentery model, were demonstrated in various studies; e.g., real-time quantitative detection of circulating and migrating individual blood and cancer cells, studies on vascular dynamics with a focus on lymphatics under normal conditions and under different interventions (e.g. lasers, drugs, nicotine), assessment of lymphatic disturbances from experimental lymphedema, monitoring cell traffic between blood and lymph systems, and high-speed imaging of cell transient deformability in flow. In particular, the obtained results demonstrated that individual cell transportation in living organisms depends on cell type (e.g., normal blood or leukemic cells), the cell’s functional state (e.g., live, apoptotic, or necrotic), and the functional status of the organism. Possible future applications, including in vivo early diagnosis and prevention of disease, monitoring immune response and apoptosis, chemo- and radio-sensitivity tests, and drug screening, are also discussed. PMID:17226898

Liver injury in hypervitaminosis A: Evidence for activation of Kupffer cell function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sim, W.L.W.

1988-01-01

The most important and novel finding of this work was enhanced liver Kupffer cell phagocytic and metabolic function by hypervitaminosis A. An animal model of hypervitaminosis A was developed in male Sprague-Dawley rats gavaged with 250,000 I.U. retinol/kg body weight/day for 3 weeks. Presence of hypervitaminosis A was indicated by characteristic changes in the fur coat, presence of brittle bones and spontaneous fractures and a significant increase in plasma and liver concentrations of retinyl palmitate while retinol levels remained the same as in controls. Hypervitaminosis A did not cause severe liver abnormalities as reflected by normal plasma glutamate pyruvate transaminasemore » activity and bilirubin. The main change was a marked increase in size of the fat or Vitamin A storing cells. Measurement of clearance from blood of indocyanine green and {sup 99m}Tc-disofenin indicated this hepatocyte function was normal. Kupffer cell phagocytic function was enhanced in hypervitaminosis A as determined by clearance from blood of {sup 99m}Tc-sulfur colloid. In vitro, there was also evidence that treatment with high doses of Vitamin A activated or enhanced Kupffer cell function. Kupffer cells from control and Vitamin A treated rats were isolated by enzymatic dispersion, purified by centrifugal elutriation, and placed in culture. Activation was indicated by (1) increased phagocytosis of {sup 51}Cr-labeled opsonized sheep red blood cells (2) enhanced release of superoxide anion and (3) enhanced production of tumor cytolytic factor by Kupffer cells from Vitamin A treated rats.« less

Method for determining properties of red blood cells

DOEpatents

Gourley, Paul L.

2001-01-01

A method for quantifying the concentration of hemoglobin in a cell, and indicia of anemia, comprises determining the wavelength of the longitudinal mode of a liquid in a laser microcavity; determining the wavelength of the fundamental transverse mode of a red blood cell in the liquid in the laser microcavity; and determining if the cell is anemic from the difference between the wavelength of the longitudinal mode and the fundamental transverse mode. In addition to measuring hemoglobin, the invention includes a method using intracavity laser spectroscopy to measure the change in spectra as a function of time for measuring the influx of water into a red blood cell and the cell's subsequent rupture.

Surfactant functionalization induces robust, differential adhesion of tumor cells and blood cells to charged nanotube-coated biomaterials under flow.

PubMed

Mitchell, Michael J; Castellanos, Carlos A; King, Michael R

2015-07-01

The metastatic spread of cancer cells from the primary tumor to distant sites leads to a poor prognosis in cancers originating from multiple organs. Increasing evidence has linked selectin-based adhesion between circulating tumor cells (CTCs) and endothelial cells of the microvasculature to metastatic dissemination, in a manner similar to leukocyte adhesion during inflammation. Functionalized biomaterial surfaces hold promise as a diagnostic tool to separate CTCs and potentially treat metastasis, utilizing antibody and selectin-mediated interactions for cell capture under flow. However, capture at high purity levels is challenged by the fact that CTCs and leukocytes both possess selectin ligands. Here, a straightforward technique to functionalize and alter the charge of naturally occurring halloysite nanotubes using surfactants is reported to induce robust, differential adhesion of tumor cells and blood cells to nanotube-coated surfaces under flow. Negatively charged sodium dodecanoate-functionalized nanotubes simultaneously enhanced tumor cell capture while negating leukocyte adhesion, both in the presence and absence of adhesion proteins, and can be utilized to isolate circulating tumor cells regardless of biomarker expression. Conversely, diminishing nanotube charge via functionalization with decyltrimethylammonium bromide both abolished tumor cell capture while promoting leukocyte adhesion. Copyright © 2015 Elsevier Ltd. All rights reserved.

Maternal Body-Mass Index and Cord Blood Circulating Endothelial Colony-Forming Cells

PubMed Central

Lin, Ruei-Zeng; Miranda, Maria L.; Vallejo-Vaz, Antonio J.; Stiefel, Pablo; Praena-Fernández, Juan M.; Bernal-Bermejo, Jose; Jimenez-Jimenez, Luis M.; Villar, Jose; Melero-Martin, Juan M.

2013-01-01

Objective Endothelial colony-forming cells (ECFCs) are a subset of circulating endothelial progenitor cells that are particularly abundant in umbilical cord blood. We sought to determine whether ECFC abundance in cord blood is associated with maternal body-mass index (BMI) in non-pathological pregnancies. Study design We measured the level of ECFCs in the cord blood of neonates (n=27) born from non-obese healthy mothers with non-pathological pregnancies and examined whether ECFC abundance correlated with maternal BMI. We also examined the effect of maternal BMI on ECFC phenotype and function using angiogenic and vasculogenic assays. Results We observed variation in ECFC abundance among subjects and found a positive correlation between pre-pregnancy maternal BMI and ECFC content (r=0.51, P=0.007), which was independent of other obstetric factors. Despite this variation, ECFC phenotype and functionality were deemed normal and highly similar between subjects with maternal BMI <25 kg/m2 and BMI between 25–30 kg/m2, including the ability to form vascular networks in vivo. Conclusions This study underlines the need to consider maternal BMI as a potential confounding factor for cord blood levels of ECFCs in future comparative studies between healthy and pathological pregnancies. Endothelial colony-forming cells (ECFCs) are a subset of progenitor cells that circulate in peripheral blood and can give rise to endothelial cells (1,2), contributing to the formation of new vasculature and the maintenance of vascular integrity (3–5). The mechanisms that regulate the abundance of these cells in vivo remain poorly understood. ECFCs are rare in adult peripheral blood (1,2,10). In contrast, there is an elevated number of these cells in fetal blood during the third trimester of pregnancy (11–13). Emerging evidence indicates that deleterious conditions during fetal life can impair ECFC content and function. For instance, offspring of diabetic mothers have been shown to have reduced number of circulating ECFCs and impaired cell functionality (14), which may contribute to the long-term cardiovascular complications. Similar observations have been reported in neonates with bronchopulmonary dysplasia (15,16). The adverse association between maternal weight and the outcome of pregnancy is well known (17,18). Epidemiologic studies have shown that cardiovascular disease may have origins during fetal development (19). Excessive maternal pre-pregnancy weight and gestational weight gain are associated with adverse cardiovascular risk factors in the offspring (20). The fetal adaptations that occur in response to changes in maternal weight during pregnancy and whether these adaptations affect the level of ECFCs is completely unknown. In this study we quantified the baseline variation in ECFC abundance and function among neonates born from non-obese healthy mothers with non-pathological pregnancies and examined whether this normal variation was associated with differences in maternal weight. PMID:24315508

CAVEOLINS AND LUNG FUNCTION

PubMed Central

Maniatis, Nikolaos A.; Chernaya, Olga; Shinin, Vasily; Minshall, Richard D.

2012-01-01

The primary function of the mammalian lung is to facilitate diffusion of oxygen to venous blood and to ventilate carbon dioxide produced by catabolic reactions within cells. However, it is also responsible for a variety of other important functions, including host defense and production of vasoactive agents to regulate not only systemic blood pressure, but also water, electrolyte and acid-base balance. Caveolin-1 is highly expressed in the majority of cell types in the lung, including epithelial, endothelial, smooth muscle, connective tissue cells, and alveolar macrophages. Deletion of caveolin-1 in these cells results in major functional aberrations, suggesting that caveolin-1 may be crucial to lung homeostasis and development. Furthermore, generation of mutant mice that under-express caveolin-1 results in severe functional distortion with phenotypes covering practically the entire spectrum of known lung diseases, including pulmonary hypertension, fibrosis, increased endothelial permeability, and immune defects. In this Chapter, we outline the current state of knowledge regarding caveolin-1-dependent regulation of pulmonary cell functions and discuss recent research findings on the role of caveolin-1 in various pulmonary disease states, including obstructive and fibrotic pulmonary vascular and inflammatory diseases. PMID:22411320

Differential Mechanisms Associated with Vascular Disrupting Action of Electrochemotherapy: Intravital Microscopy on the Level of Single Normal and Tumor Blood Vessels

PubMed Central

Markelc, Bostjan; Sersa, Gregor; Cemazar, Maja

2013-01-01

Electropermeabilization/electroporation (EP) provides a tool for the introduction of molecules into cells and tissues. In electrochemotherapy (ECT), cytotoxic drugs are introduced into cells in tumors, and nucleic acids are introduced into cells in gene electrotransfer. The normal and tumor tissue blood flow modifying effects of EP and the vascular disrupting effect of ECT in tumors have already been determined. However, differential effects between normal vs. tumor vessels, to ensure safety in the clinical application of ECT, have not been determined yet. Therefore, the aim of our study was to determine the effects of EP and ECT with bleomycin on the HT-29 human colon carcinoma tumor model and its surrounding blood vessels. The response of blood vessels to EP and ECT was monitored in real time, directly at the single blood vessel level, by in vivo optical imaging in a dorsal window chamber in SCID mice with 70 kDa fluorescently labeled dextrans. The response of tumor blood vessels to EP and ECT started to differ within the first hour. Both therapies induced a vascular lock, decreased functional vascular density (FVD) and increased the diameter of functional blood vessels within the tumor. The effects were more pronounced for ECT, which destroyed the tumor blood vessels within 24 h. Although the vasculature surrounding the tumor was affected by EP and ECT, it remained functional. The study confirms the current model of tumor blood flow modifying effects of EP and provides conclusive evidence that ECT is a vascular disrupting therapy with a specific effect on the tumor blood vessels. PMID:23555705

Functional Module Search in Protein Networks based on Semantic Similarity Improves the Analysis of Proteomics Data*

PubMed Central

Boyanova, Desislava; Nilla, Santosh; Klau, Gunnar W.; Dandekar, Thomas; Müller, Tobias; Dittrich, Marcus

2014-01-01

The continuously evolving field of proteomics produces increasing amounts of data while improving the quality of protein identifications. Albeit quantitative measurements are becoming more popular, many proteomic studies are still based on non-quantitative methods for protein identification. These studies result in potentially large sets of identified proteins, where the biological interpretation of proteins can be challenging. Systems biology develops innovative network-based methods, which allow an integrated analysis of these data. Here we present a novel approach, which combines prior knowledge of protein-protein interactions (PPI) with proteomics data using functional similarity measurements of interacting proteins. This integrated network analysis exactly identifies network modules with a maximal consistent functional similarity reflecting biological processes of the investigated cells. We validated our approach on small (H9N2 virus-infected gastric cells) and large (blood constituents) proteomic data sets. Using this novel algorithm, we identified characteristic functional modules in virus-infected cells, comprising key signaling proteins (e.g. the stress-related kinase RAF1) and demonstrate that this method allows a module-based functional characterization of cell types. Analysis of a large proteome data set of blood constituents resulted in clear separation of blood cells according to their developmental origin. A detailed investigation of the T-cell proteome further illustrates how the algorithm partitions large networks into functional subnetworks each representing specific cellular functions. These results demonstrate that the integrated network approach not only allows a detailed analysis of proteome networks but also yields a functional decomposition of complex proteomic data sets and thereby provides deeper insights into the underlying cellular processes of the investigated system. PMID:24807868

Deconvoluting Post-Transplant Immunity: Cell Subset-Specific Mapping Reveals Pathways for Activation and Expansion of Memory T, Monocytes and B Cells

PubMed Central

Grigoryev, Yevgeniy A.; Kurian, Sunil M.; Avnur, Zafi; Borie, Dominic; Deng, Jun; Campbell, Daniel; Sung, Joanna; Nikolcheva, Tania; Quinn, Anthony; Schulman, Howard; Peng, Stanford L.; Schaffer, Randolph; Fisher, Jonathan; Mondala, Tony; Head, Steven; Flechner, Stuart M.; Kantor, Aaron B.; Marsh, Christopher; Salomon, Daniel R.

2010-01-01

A major challenge for the field of transplantation is the lack of understanding of genomic and molecular drivers of early post-transplant immunity. The early immune response creates a complex milieu that determines the course of ensuing immune events and the ultimate outcome of the transplant. The objective of the current study was to mechanistically deconvolute the early immune response by purifying and profiling the constituent cell subsets of the peripheral blood. We employed genome-wide profiling of whole blood and purified CD4, CD8, B cells and monocytes in tandem with high-throughput laser-scanning cytometry in 10 kidney transplants sampled serially pre-transplant, 1, 2, 4, 8 and 12 weeks. Cytometry confirmed early cell subset depletion by antibody induction and immunosuppression. Multiple markers revealed the activation and proliferative expansion of CD45RO+CD62L− effector memory CD4/CD8 T cells as well as progressive activation of monocytes and B cells. Next, we mechanistically deconvoluted early post-transplant immunity by serial monitoring of whole blood using DNA microarrays. Parallel analysis of cell subset-specific gene expression revealed a unique spectrum of time-dependent changes and functional pathways. Gene expression profiling results were validated with 157 different probesets matching all 65 antigens detected by cytometry. Thus, serial blood cell monitoring reflects the profound changes in blood cell composition and immune activation early post-transplant. Each cell subset reveals distinct pathways and functional programs. These changes illuminate a complex, early phase of immunity and inflammation that includes activation and proliferative expansion of the memory effector and regulatory cells that may determine the phenotype and outcome of the kidney transplant. PMID:20976225

Deconvoluting post-transplant immunity: cell subset-specific mapping reveals pathways for activation and expansion of memory T, monocytes and B cells.

PubMed

Grigoryev, Yevgeniy A; Kurian, Sunil M; Avnur, Zafi; Borie, Dominic; Deng, Jun; Campbell, Daniel; Sung, Joanna; Nikolcheva, Tania; Quinn, Anthony; Schulman, Howard; Peng, Stanford L; Schaffer, Randolph; Fisher, Jonathan; Mondala, Tony; Head, Steven; Flechner, Stuart M; Kantor, Aaron B; Marsh, Christopher; Salomon, Daniel R

2010-10-14

A major challenge for the field of transplantation is the lack of understanding of genomic and molecular drivers of early post-transplant immunity. The early immune response creates a complex milieu that determines the course of ensuing immune events and the ultimate outcome of the transplant. The objective of the current study was to mechanistically deconvolute the early immune response by purifying and profiling the constituent cell subsets of the peripheral blood. We employed genome-wide profiling of whole blood and purified CD4, CD8, B cells and monocytes in tandem with high-throughput laser-scanning cytometry in 10 kidney transplants sampled serially pre-transplant, 1, 2, 4, 8 and 12 weeks. Cytometry confirmed early cell subset depletion by antibody induction and immunosuppression. Multiple markers revealed the activation and proliferative expansion of CD45RO(+)CD62L(-) effector memory CD4/CD8 T cells as well as progressive activation of monocytes and B cells. Next, we mechanistically deconvoluted early post-transplant immunity by serial monitoring of whole blood using DNA microarrays. Parallel analysis of cell subset-specific gene expression revealed a unique spectrum of time-dependent changes and functional pathways. Gene expression profiling results were validated with 157 different probesets matching all 65 antigens detected by cytometry. Thus, serial blood cell monitoring reflects the profound changes in blood cell composition and immune activation early post-transplant. Each cell subset reveals distinct pathways and functional programs. These changes illuminate a complex, early phase of immunity and inflammation that includes activation and proliferative expansion of the memory effector and regulatory cells that may determine the phenotype and outcome of the kidney transplant.

Steady state peripheral blood provides cells with functional and metabolic characteristics of real hematopoietic stem cells.

PubMed

Bourdieu, Antonin; Avalon, Maryse; Lapostolle, Véronique; Ismail, Sadek; Mombled, Margaux; Debeissat, Christelle; Guérinet, Marianne; Duchez, Pascale; Chevaleyre, Jean; Vlaski-Lafarge, Marija; Villacreces, Arnaud; Praloran, Vincent; Ivanovic, Zoran; Brunet de la Grange, Philippe

2018-01-01

Hematopoietic stem cells (HSCs), which are located in the bone marrow, also circulate in cord and peripheral blood. Despite high availability, HSCs from steady state peripheral blood (SSPB) are little known and not used for research or cell therapy. We thus aimed to characterize and select HSCs from SSPB by a direct approach with a view to delineating their main functional and metabolic properties and the mechanisms responsible for their maintenance. We chose to work on Side Population (SP) cells which are highly enriched in HSCs in mouse, human bone marrow, and cord blood. However, no SP cells from SSBP have as yet been characterized. Here we showed that SP cells from SSPB exhibited a higher proliferative capacity and generated more clonogenic progenitors than non-SP cells in vitro. Furthermore, xenotransplantation studies on immunodeficient mice demonstrated that SP cells are up to 45 times more enriched in cells with engraftment capacity than non-SP cells. From a cell regulation point of view, we showed that SP activity depended on O 2 concentrations close to those found in HSC niches, an effect which is dependent on both hypoxia-induced factors HIF-1α and HIF-2α. Moreover SP cells displayed a reduced mitochondrial mass and, in particular, a lower mitochondrial activity compared to non-SP cells, while they exhibited a similar level of glucose incorporation. These results provided evidence that SP cells from SSPB displayed properties of very primitive cells and HSC, thus rendering them an interesting model for research and cell therapy. © 2017 Wiley Periodicals, Inc.

Phenotypic and Functional Characterization of Peripheral Blood Lymphocytes from Various Age- and Sex-Specific Groups of Owl Monkeys (Aotus nancymaae).

PubMed

Nehete, Pramod N; Nehete, Bharti P; Chitta, Sriram; Williams, Lawrence E; Abee, Christian R

2017-02-01

Owl monkeys (Aotus nancymaae) are New World NHP that serve an important role in vaccine development and as a model for human disease conditions such as malaria. Despite the past contributions of this animal model, limited information is available about the phenotype and functional properties of peripheral blood lymphocytes in reference to sex and age. Using a panel of human antibodies and a set of standardized human immune assays, we identified and characterized various peripheral blood lymphocyte subsets, evaluated the immune functions of T cells, and analyzed cytokines relative to sex and age in healthy owl monkeys. We noted age- and sex-dependent changes in CD28+ (an essential T cell costimulatory molecule) and CD95+ (an apoptotic surface marker) T cells and various levels of cytokines in the plasma. In immune assays of freshly isolated peripheral blood mononuclear cells, IFNγ and perforin responses were significantly higher in female than in male monkeys and in young adults than in juvenile and geriatric groups, despite similar lymphocyte (particularly T cell) populations in these groups. Our current findings may be useful in exploring Aotus monkeys as a model system for the study of aging, susceptibility to infectious diseases, and age-associated differences in vaccine efficacy, and other challenges particular to pediatric and geriatric patients.

Thiol-Reactive Star Polymers Display Enhanced Association with Distinct Human Blood Components.

PubMed

Glass, Joshua J; Li, Yang; De Rose, Robert; Johnston, Angus P R; Czuba, Ewa I; Khor, Song Yang; Quinn, John F; Whittaker, Michael R; Davis, Thomas P; Kent, Stephen J

2017-04-12

Directing nanoparticles to specific cell types using nonantibody-based methods is of increasing interest. Thiol-reactive nanoparticles can enhance the efficiency of cargo delivery into specific cells through interactions with cell-surface proteins. However, studies to date using this technique have been largely limited to immortalized cell lines or rodents, and the utility of this technology on primary human cells is unknown. Herein, we used RAFT polymerization to prepare pyridyl disulfide (PDS)-functionalized star polymers with a methoxy-poly(ethylene glycol) brush corona and a fluorescently labeled cross-linked core using an arm-first method. PDS star polymers were examined for their interaction with primary human blood components: six separate white blood cell subsets, as well as red blood cells and platelets. Compared with control star polymers, thiol-reactive nanoparticles displayed enhanced association with white blood cells at 37 °C, particularly the phagocytic monocyte, granulocyte, and dendritic cell subsets. Platelets associated with more PDS than control nanoparticles at both 37 °C and on ice, but they were not activated in the duration examined. Association with red blood cells was minor but still enhanced with PDS nanoparticles. Thiol-reactive nanoparticles represent a useful strategy to target primary human immune cell subsets for improved nanoparticle delivery.

Sensitive capture of circulating tumour cells by functionalized graphene oxide nanosheets

NASA Astrophysics Data System (ADS)

Yoon, Hyeun Joong; Kim, Tae Hyun; Zhang, Zhuo; Azizi, Ebrahim; Pham, Trinh M.; Paoletti, Costanza; Lin, Jules; Ramnath, Nithya; Wicha, Max S.; Hayes, Daniel F.; Simeone, Diane M.; Nagrath, Sunitha

2013-10-01

The spread of cancer throughout the body is driven by circulating tumour cells (CTCs). These cells detach from the primary tumour and move from the bloodstream to a new site of subsequent tumour growth. They also carry information about the primary tumour and have the potential to be valuable biomarkers for disease diagnosis and progression, and for the molecular characterization of certain biological properties of the tumour. However, the limited sensitivity and specificity of current methods for measuring and studying these cells in patient blood samples prevents the realization of their full clinical potential. The use of microfluidic devices is a promising method for isolating CTCs. However, the devices are reliant on three-dimensional structures, which limits further characterization and expansion of cells on the chip. Here we demonstrate an effective approach to isolating CTCs from blood samples of pancreatic, breast and lung cancer patients, by using functionalized graphene oxide nanosheets on a patterned gold surface. CTCs were captured with high sensitivity at a low concentration of target cells (73 +/- 32.4% at 3-5 cells per ml blood).

Effect of Flow on Gene Regulation in Smooth Muscle Cells and Macromolecular Transport Across Endothelial Cell Monolayers

NASA Technical Reports Server (NTRS)

McIntire, Larry V.; Wagner, John E.; Papadaki, Maria; Whitson, Peggy A.; Eskin, Suzanne G.

1996-01-01

Endothelial cells line all of the vessels of the circulatory system, providing a non-thrombogenic conduit for blood flow; they regulate many complex functions in the vasculature, such as coagulation, fibrinolysis, platelet aggregation, vessel tone and growth, and leukocyte traffic; and they form the principal barrier to transport of substances between the blood and the surrounding tissue space. The permeability of endothelial cell changes with environmental stimuli; shear stress, in particular, applied either in vivo, or in vitro, induces changes in protein expression and secretion of vasoactive factors by endothelial cells. The ability to study the effects of shear on the macromolecular permeability of the cerebral vasculature is particularly important, since in no other place is the barrier function of the endothelium more important than in the brain. The endothelial cells of this organ have developed special barrier properties that keep the cerebral system from experiencing any drastic change in composition; together with glial cells, they form the blood brain barrier (BBB). We have studied the effect of flow on bovine BBB using flow chambers and tissue culture systems.

Direct measurement of IgM-Antigen interaction energy on individual red blood cells.

PubMed

Yeow, Natasha; Tabor, Rico F; Garnier, Gil

2017-07-01

Most blood grouping tests rely on the principle of red blood cells (RBCs) agglutination. Agglutination is triggered by the binding of specific blood grouping antibodies to the corresponding RBC surface antigen on multiple cells. The interaction energies between blood grouping antibodies and antigens have been poorly defined in immunohaematology. Here for the first time, we functionalized atomic force microscope (AFM) cantilevers with the IgM form of blood grouping antibodies to probe populations of individual RBCs of different groups under physiological conditions. The force-mapping mode of AFM allowed us to measure specific antibody - antigen interactions, and simultaneously localize and quantify antigen sites on the scanned cell surface. This study provides a new insight of the interactions between IgM antibodies and its corresponding antigen. The technique and information can be translated to develop better blood typing diagnostics and optimize target-specific drug delivery for medical applications. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

Microparticles Engineered to Highly Express Peroxisome Proliferator-Activated Receptor-γ Decreased Inflammatory Mediator Production and Increased Adhesion of Recipient Monocytes

PubMed Central

Sahler, Julie; Woeller, Collynn F.; Phipps, Richard P.

2014-01-01

Circulating blood microparticles are submicron vesicles released primarily by megakaryocytes and platelets that act as transcellular communicators. Inflammatory conditions exhibit elevated blood microparticle numbers compared to healthy conditions. Direct functional consequences of microparticle composition, especially internal composition, on recipient cells are poorly understood. Our objective was to evaluate if microparticle composition could impact the function of recipient cells, particularly during inflammatory provocation. We therefore engineered the composition of megakaryocyte culture-derived microparticles to generate distinct microparticle populations that were given to human monocytes to assay for influences recipient cell function. Herein, we tested the responses of monocytes exposed to either control microparticles or microparticles that contain the anti-inflammatory transcription factor, peroxisome proliferator-activated receptor-γ (PPARγ). In order to normalize relative microparticle abundance from two microparticle populations, we implemented a novel approach that utilizes a Nanodrop Spectrophotometer to assay for microparticle density rather than concentration. We found that when given to peripheral blood mononuclear cells, microparticles were preferentially internalized by CD11b+ cells, and furthermore, microparticle composition had a profound functional impact on recipient monocytes. Specifically, microparticles containing PPARγ reduced activated monocyte production of the proinflammatory cytokines interleukin-8 and monocyte chemotactic protein-1 compared to activated monocytes exposed to control microparticles. Additionally, treatment with PPARγ microparticles greatly increased monocyte cell adherence. This change in morphology occurred simultaneously with increased production of the key extracellular matrix protein, fibronectin and increased expression of the fibronectin-binding integrin, ITGA5. PPARγ microparticles also changed monocyte mRNA levels of several genes including those under PPARγ control. Overall, the delivery of PPARγ from microparticles to human monocytes influenced gene expression, decreased inflammatory mediator production and increased monocyte adherence. These results support the concept that the composition of blood microparticles has a profound impact on the function of cells with which they interact, and likely plays a role in vascular inflammation. PMID:25426628

Microparticles engineered to highly express peroxisome proliferator-activated receptor-γ decreased inflammatory mediator production and increased adhesion of recipient monocytes.

PubMed

Sahler, Julie; Woeller, Collynn F; Phipps, Richard P

2014-01-01

Circulating blood microparticles are submicron vesicles released primarily by megakaryocytes and platelets that act as transcellular communicators. Inflammatory conditions exhibit elevated blood microparticle numbers compared to healthy conditions. Direct functional consequences of microparticle composition, especially internal composition, on recipient cells are poorly understood. Our objective was to evaluate if microparticle composition could impact the function of recipient cells, particularly during inflammatory provocation. We therefore engineered the composition of megakaryocyte culture-derived microparticles to generate distinct microparticle populations that were given to human monocytes to assay for influences recipient cell function. Herein, we tested the responses of monocytes exposed to either control microparticles or microparticles that contain the anti-inflammatory transcription factor, peroxisome proliferator-activated receptor-γ (PPARγ). In order to normalize relative microparticle abundance from two microparticle populations, we implemented a novel approach that utilizes a Nanodrop Spectrophotometer to assay for microparticle density rather than concentration. We found that when given to peripheral blood mononuclear cells, microparticles were preferentially internalized by CD11b+ cells, and furthermore, microparticle composition had a profound functional impact on recipient monocytes. Specifically, microparticles containing PPARγ reduced activated monocyte production of the proinflammatory cytokines interleukin-8 and monocyte chemotactic protein-1 compared to activated monocytes exposed to control microparticles. Additionally, treatment with PPARγ microparticles greatly increased monocyte cell adherence. This change in morphology occurred simultaneously with increased production of the key extracellular matrix protein, fibronectin and increased expression of the fibronectin-binding integrin, ITGA5. PPARγ microparticles also changed monocyte mRNA levels of several genes including those under PPARγ control. Overall, the delivery of PPARγ from microparticles to human monocytes influenced gene expression, decreased inflammatory mediator production and increased monocyte adherence. These results support the concept that the composition of blood microparticles has a profound impact on the function of cells with which they interact, and likely plays a role in vascular inflammation.

T-cell receptor transfer for boosting HIV-1-specific T-cell immunity in HIV-1-infected patients.

PubMed

Mummert, Christiane; Hofmann, Christian; Hückelhoven, Angela G; Bergmann, Silke; Mueller-Schmucker, Sandra M; Harrer, Ellen G; Dörrie, Jan; Schaft, Niels; Harrer, Thomas

2016-09-10

Strategies to cure HIV-1 infection require the eradication of viral reservoirs. An innovative approach for boosting the cytotoxic T-lymphocyte response is the transfer of T-cell receptors (TCRs). Previously, we have shown that electroporation of TCR-encoding mRNA is able to reprogram CD8 T cells derived from healthy donors. So far, it is unknown whether the transfer of HIV-1-specific TCRs is capable to reprogram CD8 T cells of HIV-1-infected patients. To assess the efficiency of TCR-transfer by mRNA electroporation and the functionality of reprogramed T cells in HIV-1-infected patients, we performed an in-vitro analysis of TCR-transfer into T cells from HIV-1-infected patients in various stages of disease and from healthy controls. Peripheral blood mononuclear cells from 16 HIV-1-infected patients (nine HLA-A02-positive, seven HLA-A02-negative) and from five healthy controls were electroporated with mRNA-constructs encoding TCRs specific for the HLA-A02/HIV-1-gag p17 epitope SLYNTVATL (SL9). Functionality of the TCRs was measured by γIFN-ELISpot assays. SL9/TCR transfection into peripheral blood mononuclear cells from both HLA-A02-positive and HLA-A02-negative HIV-1-infected patients and from healthy blood donors reprogramed T cells for recognition of SL9-presenting HLA-A02-positive cells in γIFN-ELISpot assays. SL9/TCR-transfer into T cells from an immunodeficient AIDS patient could induce recognition of SL9-expressing target cells only after reversion of T-cell dysfunction by antiretroviral therapy. The transfer of HIV-1-p17-specific TCRs into T cells is functional both in HIV-1-infected patients as well as in healthy blood donors. TCR-transfer is a promising method to boost the immune system against HIV-1.

Elevated levels of polymorphonuclear myeloid-derived suppressor cells in patients with glioblastoma highly express S100A8/9 and arginase and suppress T cell function

PubMed Central

Gielen, Paul R.; Schulte, Barbara M.; Kers-Rebel, Esther D.; Verrijp, Kiek; Bossman, Sandra A.J.F.H.; ter Laan, Mark; Wesseling, Pieter

2016-01-01

Background Gliomas are primary brain tumors that are associated with a poor prognosis. The introduction of new treatment modalities (including immunotherapy) for these neoplasms in the last 3 decades has resulted in only limited improvement in survival. Gliomas are known to create an immunosuppressive microenvironment that hampers the efficacy of (immuno)therapy. One component of this immunosuppressive environment is the myeloid-derived suppressor cell (MDSC). Methods We set out to analyze the presence and activation state of MDSCs in blood (n = 41) and tumor (n = 20) of glioma patients by measuring S100A8/9 and arginase using flow cytometry and qPCR. Inhibition of T cell proliferation and cytokine production after stimulation with anti-CD3/anti-CD28 coated beads was used to measure in vitro MDSC suppression capacity. Results We report a trend toward a tumor grade-dependent increase of both monocytic (M-) and polymorphonuclear (PMN-) MDSC subpopulations in the blood of patients with glioma. M-MDSCs of glioma patients have increased levels of intracellular S100A8/9 compared with M-MDSCs in healthy controls (HCs). Glioma patients also have increased S100A8/9 serum levels, which correlates with increased arginase activity in serum. PMN-MDSCs in both blood and tumor tissue demonstrated high expression of arginase. Furthermore, we assessed blood-derived PMN-MDSC function and showed that these cells have potent T cell suppressive function in vitro. Conclusions These data indicate a tumor grade-dependent increase of MDSCs in the blood of patients with a glioma. These MDSCs exhibit an increased activation state compared with MDSCs in HCs, independent of tumor grade. PMID:27006175

[Effects of Chinese herbal compound for supplementing qi and activating blood circulation on actin, Cx43 expressions and gap junctional intercellular communication functions of myocardial cells in patients with Coxsackie virus B 3 viral myocarditis].

PubMed

Zhang, Ming-xue; He, Wei; Gu, Ping

2010-08-01

To observe the effect of Chinese herbal compound for supplementing qi and activating blood circulation (CHC) on the gap junctional intercellular communication (GJIC) function of myocardial cells in patients with Coxsackie virus B 3 (CVB3) viral myocarditis. Expressions of actin and connexin43 (Cx43) in myocardial cells of patients arranged in three groups (the normal control group, the viral infected group and the CHC treated group) were detected by immunohistochemical method; the fluorescence photobleaching recovery rate of cells was detected by laser scanning confocal microscope. As compared with the viral infected group, the expressions of actin and Cx43 were increased and the GJIC function was improved in the CHC treated group. CHC could antagonize viral injury on skeleton protein, and repair the structure of gap junction channel to improve the GJIC function of myocardial cells after being attacked by CVB3.

The albumin-exendin-4 recombinant protein E2HSA improves glycemic control and β-cell function in spontaneous diabetic KKAy mice.

PubMed

Li, Caina; Hou, Shaocong; Liu, Shuainan; Huan, Yi; Sun, Sujuan; Liu, Quan; Shen, Zhufang

2017-06-19

E2HSA is a genetic fusion protein that consists of two tandem exendin-4 molecules that are covalently bonded to recombinant human serum albumin via a peptide linker. Previous studies have demonstrated that E2HSA significantly decreased blood glucose levels, improved β-cell function and promoted β-cell proliferation in diabetic db/dB mice. This study aimed to evaluate the benefits of E2HSA on glucose and lipid metabolism in a spontaneous diabetes animal model, KKAy mice. E2HSA was acutely administered at doses of 1, 3 and 9 mg/kg by subcutaneous injection in diabetic KKAy mice with exendin-4 (2 μg/kg) as a positive reference, and then the non-fasting blood glucose and food intake levels were dynamically monitored. In addition, different doses of E2HSA were injected once daily, as well as with exendin-4 twice daily, for 7 weeks to evaluate the effect on glucose and lipid metabolism, as well as the body weight, food and water intake. Single injection of E2HSA decreased non-fasting blood glucose and food intake levels in a dose-dependent manner for 4 days and 2 days, respectively. Repeated injections with E2HSA significantly decreased variations in blood glucose levels with a reduction of HbA1c levels by 1.6% at a 9 mg/kg dose, simultaneously increased fasting blood insulin levels, inhibited fasting blood glucagon levels, improved the impaired oral glucose tolerance and enhanced glucose infusion rate, which is the gold standard for evaluating β-cell function. Moreover, repeated injections with E2HSA also ameliorated the dyslipidemia and reduced body weight, food and water intake in diabetic KKAy mice. E2HSA significantly reduced blood glucose levels over a prolonged duration, enhanced β-cell function, and ameliorated dyslipidemia and obesity in diabetic KKAy mice. Thus, E2HSA may be a new candidate for the treatment of type 2 diabetes.

Effect of allo- and xenotransplantation of embryonic nervous tissue and umbilical cord blood-derived stem cells on structural and functional state of cerebral cortex of albino rats in posttraumatic period.

PubMed

Ereniev, S I; Semchenko, V V; Sysheva, E V; Bogdashin, I V; Shapovalova, V V; Khizhnyak, A S; Gasanenko, L N

2005-11-01

Comparative study of the structural and functional state of cerebral cortex of adult albino rats after intracerebral allo- and xenotransplantation of embryonic nervous tissue and intravenous injection of umbilical cord blood-derived stem cells at different terms after diffuse-focal cerebral trauma revealed the best cerebroprotective effect on day 7 of posttraumatic period in animals receiving embryonic nervous tissue.

[The phylogenetic theory of pathology. The arterial hypertension--a test of metabolic disorder. The biological basis of damage of target organs (a lecture)].

PubMed

Titov, V N

2013-05-01

The increase of blood tension is a diagnostic test of disorders of homeostasis, trophology, endoecology and adaptation in paracrine regulated coenosis of cells. This conditions results in disorder of microcirculation in the distal section of arterial race and in compensatory increase of blood tension in its proximal section. The increase of blood tension disturbs the function of paracrine coenosis of cells which have one's own system of hemo- and hydrodynamics such as brain with system of spinal liquor and kidneys with local pool of primary urine. They counteract the rise of blood tension and activate local, humoral system of renin-angiotensin-II increasing peripheral resistance to blood flow. At that, the compensatory blood tension becomes even higher. The aldosterone and natriuretic peptides are functional synergists. So, they preserve and excrete ions of Na+ and support the stability of unified pool of intercellular medium ("Inner Ocean" of organism) where all cells live. The parameters of this pool are limited most strictly in vivo. If at the level of nephron the conditions are formed that can alter the parameters of unified pool of intercellular medium the vasomotor center rises blood tension from the level of organism "forcing" nephrons to re-establish the parameters of this pool and normalize the biological functions and biological reactions. The blood pressure increase under pathology of kidneys is caused because of pathological compensation at the level of organism mediated by vegetal nervous system and dictated by necessity to preserve the parameters of inner medium of organism.

Three-dimensional biomimetic vascular model reveals a RhoA, Rac1, and N-cadherin balance in mural cell-endothelial cell-regulated barrier function.

PubMed

Alimperti, Stella; Mirabella, Teodelinda; Bajaj, Varnica; Polacheck, William; Pirone, Dana M; Duffield, Jeremy; Eyckmans, Jeroen; Assoian, Richard K; Chen, Christopher S

2017-08-15

The integrity of the endothelial barrier between circulating blood and tissue is important for blood vessel function and, ultimately, for organ homeostasis. Here, we developed a vessel-on-a-chip with perfused endothelialized channels lined with human bone marrow stromal cells, which adopt a mural cell-like phenotype that recapitulates barrier function of the vasculature. In this model, barrier function is compromised upon exposure to inflammatory factors such as LPS, thrombin, and TNFα, as has been observed in vivo. Interestingly, we observed a rapid physical withdrawal of mural cells from the endothelium that was accompanied by an inhibition of endogenous Rac1 activity and increase in RhoA activity in the mural cells themselves upon inflammation. Using a system to chemically induce activity in exogenously expressed Rac1 or RhoA within minutes of stimulation, we demonstrated RhoA activation induced loss of mural cell coverage on the endothelium and reduced endothelial barrier function, and this effect was abrogated when Rac1 was simultaneously activated. We further showed that N -cadherin expression in mural cells plays a key role in barrier function, as CRISPR-mediated knockout of N -cadherin in the mural cells led to loss of barrier function, and overexpression of N -cadherin in CHO cells promoted barrier function. In summary, this bicellular model demonstrates the continuous and rapid modulation of adhesive interactions between endothelial and mural cells and its impact on vascular barrier function and highlights an in vitro platform to study the biology of perivascular-endothelial interactions.

Stage-dependency of apoptosis and the blood-testis barrier in the dogfish shark (Squalus acanthias): cadmium-induced changes as assessed by vital fluorescence techniques.

PubMed

McClusky, Leon M

2006-09-01

Naturally occurring heavy metals and synthetic compounds are potentially harmful for testicular function but evidence linking heavy metal exposure to reduced semen parameters is inconclusive. Elucidation of the exact stage at which the toxicant interferes with spermatogenesis is difficult because the various germ cell stages may have different sensitivities to any given toxicant, germ cell development is influenced by supporting testicular somatic cells and the presence of inter-Sertoli cell tight junctions create a blood-testis barrier, sequestering meiotic and postmeiotic germ cells in a special microenvironment. Sharks such as Squalus acanthias provide a suitable model for studying aspects of vertebrate spermatogenosis because of their unique features: spermatogenesis takes place within spermatocysts and relies mainly on Sertoli cells for somatic cell support; spermatocysts are linearly arranged in a maturational order across the diameter of the elongated testis; spermatocysts containing germ cells at different stages of development are topographically separated, resulting in visible zonation in testicular cross sections. We have used the vital dye acridine orange and a novel fluorescence staining technique to study this model to determine (1) the efficacy of these methods in assays of apoptosis and blood-testis barrier function, (2) the sensitivity of the various spermatogonial generations in Squalus to cadmium (as an illustrative spermatotoxicant) and (3) the way that cadmium might affect more mature spermatogenic stages and other physiological processes in the testis. Our results show that cadmium targets early spermatogenic stages, where it specifically activates a cell death program in susceptible (mature) spermatogonial clones, and negatively affects blood-testis barrier function. Since other parameters are relatively unaffected by cadmium, the effects of this toxicant on apoptosis are presumably process-specific and not attributable to general toxicity.

Reversible changes in pancreatic islet structure and function produced by elevated blood glucose

PubMed Central

Brereton, Melissa F.; Iberl, Michaela; Shimomura, Kenju; Zhang, Quan; Adriaenssens, Alice E.; Proks, Peter; Spiliotis, Ioannis I.; Dace, William; Mattis, Katia K.; Ramracheya, Reshma; Gribble, Fiona M.; Reimann, Frank; Clark, Anne; Rorsman, Patrik; Ashcroft, Frances M.

2014-01-01

Diabetes is characterized by hyperglycaemia due to impaired insulin secretion and aberrant glucagon secretion resulting from changes in pancreatic islet cell function and/or mass. The extent to which hyperglycaemia per se underlies these alterations remains poorly understood. Here we show that β-cell-specific expression of a human activating KATP channel mutation in adult mice leads to rapid diabetes and marked alterations in islet morphology, ultrastructure and gene expression. Chronic hyperglycaemia is associated with a dramatic reduction in insulin-positive cells and an increase in glucagon-positive cells in islets, without alterations in cell turnover. Furthermore, some β-cells begin expressing glucagon, whilst retaining many β-cell characteristics. Hyperglycaemia, rather than KATP channel activation, underlies these changes, as they are prevented by insulin therapy and fully reversed by sulphonylureas. Our data suggest that many changes in islet structure and function associated with diabetes are attributable to hyperglycaemia alone and are reversed when blood glucose is normalized. PMID:25145789

EFFECT OF RESIDUAL SPLENIC FUNCTION AND FOLATE LEVELS ON THE FREQUENCY OF MICRONUCLEATED RED BLOOD CELLS IN SPLENECTOMIZED HUMANS

EPA Science Inventory

Rates of micronucleated erythrocytes in the peripheral blood of splenectomized individuals can be used as an index of genetic damage to erythrocyte recursor cells in the bone marrow. his is in contrast to non-splenectomized humans, whose micronucleated erythrocytes are removed by...

[Basic studies on oral administration of lentinan (I)--influence on lymphocyte subsets in peripheral venous blood].

PubMed

Hanaue, H; Tokuda, Y; Machimura, T; Tsukui, M; Mizutani, K; Huang, C M; Kamijoh, A; Kondo, Y; Ogoshi, K; Makuuchi, H

1989-08-20

The effect of oral administration of lentinan (LTN), a biological response modifier, in the control of systemic immune function was studied in 6-week old male Wistar-Imamichi SPF rats. In the LTN group, 1 mg LTN dissolved in 1 ml physiological saline was administration forcibly into the stomach twice weekly. Physiological saline alone was administered in a similar fashion to the control group. Blood samples were obtained prior to and after four and eight weeks of administration. White blood cells and lymphocyte counts were obtained and lymphocyte subsets were measured using monoclonal antibodies W3/13, W3/25 and 0 X 8 (Sera-Lab), and a laser flow cytometry system (Orthospectrum III, Orthodiagnostic System). The T cell ratio, helper/inducer T (Th) cell ratio, and suppressor/cytotoxic T (Ts) cell ratio were measured. The peripheral white blood cell count and lymphocyte count were not significantly different between the control and LTN groups. After four weeks of LTN administration, however, the LTN group showed a significantly higher T cell ratio, Th cell ratio and Th/Ts cell ratio than did the control group, and the Ts cell ratio was significantly lower. In the groups undergoing administration for eight weeks, no difference was noted in the lymphocyte subsets between the two groups. Oral administration of LTN apparently modulates the systemic immune function through T cell stimulation, especially Th cells, but continued administration may induce a tolerance to the effect of LTN.

Plants and Photosynthesis: Level III, Unit 3, Lesson 1; The Human Digestive System: Lesson 2; Functions of the Blood: Lesson 3; Human Circulation and Respiration: Lesson 4; Reproduction of a Single Cell: Lesson 5; Reproduction by Male and Female Cells: Lesson 6; The Human Reproductive System: Lesson 7; Genetics and Heredity: Lesson 8; The Nervous System: Lesson 9; The Glandular System: Lesson 10. Advanced General Education Program. A High School Self-Study Program.

ERIC Educational Resources Information Center

Manpower Administration (DOL), Washington, DC. Job Corps.

This self-study program for the high-school level contains lessons in the following subjects: Plants and Photosynthesis; The Human Digestive System; Functions of the Blood; Human Circulation and Respiration; Reproduction of a Single Cell; Reproduction by Male and Female Cells; The Human Reproductive System; Genetics and Heredity; The Nervous…

Menstrual Blood as a Potential Source of Endometrial Derived CD3+ T Cells

PubMed Central

Sabbaj, Steffanie; Hel, Zdenek; Richter, Holly E.; Mestecky, Jiri; Goepfert, Paul A.

2011-01-01

Studies of T cell-mediated immunity in the human female genital tract have been problematic due to difficulties associated with the collection of mucosal samples. Consequently, most studies rely on biopsies from the lower female genital tract or remnant tissue from hysterectomies. Availability of samples from healthy women is limited, as most studies are carried out in women with underlying pathologies. Menstruation is the cyclical sloughing off of endometrial tissue, and thus it should be a source of endometrial cells without the need for a biopsy. We isolated and phenotyped T cells from menstrual and peripheral blood and from endometrial biopsy-derived tissue from healthy women to determine the types of T cells present in this compartment. Our data demonstrated that T cells isolated from menstrual blood are a heterogeneous population of cells with markers reminiscent of blood and mucosal cells as well as unique phenotypes not represented in either compartment. T cells isolated from menstrual blood expressed increased levels of HLA-DR, αEβ7 and CXCR4 and reduced levels of CD62L relative to peripheral blood. Menstrual blood CD4+ T cells were enriched for cells expressing both CCR7 and CD45RA, markers identifying naïve T cells and were functional as determined by antigen-specific intracellular cytokine production assays. These data may open new avenues of investigation for cell mediated immune studies involving the female reproductive tract without the need for biopsies. PMID:22174921

Involvement of specific macrophage-lineage cells surrounding arterioles in barrier and scavenger function in brain cortex.

PubMed Central

Mato, M; Ookawara, S; Sakamoto, A; Aikawa, E; Ogawa, T; Mitsuhashi, U; Masuzawa, T; Suzuki, H; Honda, M; Yazaki, Y; Watanabe, E; Luoma, J; Yla-Herttuala, S; Fraser, I; Gordon, S; Kodama, T

1996-01-01

The transport of solutes between blood and brain is regulated by a specific barrier. Capillary endothelial cells of brain are known to mediate barrier function and facilitate transport. Here we report that specific cells surrounding arterioles, known as Mato's fluorescent granular perithelial (FGP) cells or perivascular microglial cells, contribute to the barrier function. Immunohistochemical and in situ hybridization studies indicate that, in normal brain cortex, type I and type II macrophage scavenger receptors are expressed only in FGP/perivascular microglial cells, and surface markers of macrophage lineage are also detected on them. These cells mediate the uptake of macromolecules, including modified low density lipoprotein, horseradish peroxidase, and ferritin injected either into the blood or into the cerebral ventricles. Accumulation of scavenged materials with aging or after the administration of a high-fat diet results in the formation of honeycomb-like foam cells and the narrowing of the lumen of arterioles in the brain cortex. These results indicate involvement of FGP/perivascular microglial cells in the barrier and scavenger functions in the central nervous system. Images Fig. 1 Fig. 2 Fig. 4 Fig. 5 Fig. 6 PMID:8622926

ICAM-1 (CD54) expression on B lymphocytes is associated with their costimulatory function and can be increased by coactivation with IL-1 and IL-7.

PubMed

Dennig, D; Lacerda, J; Yan, Y; Gasparetto, C; O'Reilly, R J

1994-07-01

Recent studies have demonstrated that acute lymphoblastic leukemia-derived pre-B cell lines are deficient in their costimulatory function for T cell proliferation in response to the mitogen Con A and the superantigens TSST-1 and SEB. Stimulation of these pre-B cells with IL-7 increased their costimulatory function which involved the B7/CD28 pathway. In the present study, we stimulated T cells with Con A, TSST-1, and SEB in the presence of peripheral blood B lineage cells that do not constitutively express B7/BB1 on their surface and investigated whether their costimulatory function could also be enhanced by IL-7. We found that, in the presence of IL-1, stimulation with IL-7 increased the costimulatory function of B cells and their surface expression level of ICAM-1 (CD54). We then investigated whether costimulatory B cell function could be inhibited by blocking the ICAM-1/LFA-1 pathway. Addition of anti-ICAM-1 mAb to the coculture of T and B cells inhibited T cell proliferation by approximately 20%. In contrast, addition of anti-LFA-1 beta (CD18) mAb, directed against the T cell ligand of ICAM-1, inhibited T cell proliferation almost completely. To determine the role of ICAM-1 in costimulatory B cell function, we sorted B cells into ICAM-1low-and ICAM-1high-expressing populations. We found that B cells expressing high levels of surface ICAM-1 elicited significantly higher T cell responses than those with low levels, suggesting that the expression level of ICAM-1 on peripheral blood B cells correlates with their costimulatory function.

[Changes of insulin resistance and islet beta cells function in subjects with high-normal blood pressure].

PubMed

Xu, Ling-Ling; Xiang, Hong-Ding; Zhang, Li-Hong; Chen, Wei; Fang, Jing-Hui

2009-08-01

To investigate the changes of insulin resistance and islet beta cells function in subjects with euglycemia and high-normal blood pressure. Total 423 subjects were divided into normal blood pressure group and high-normal blood pressure group. Body height, weight, waist and hip circumference, and biochemical data were measured. Homeostasis model assessment of insulin resistance (HOMA-IR), insulin sensitivity index (ISI)-composite, and first-phase (1 PH) Stumvoll index were calculated. Results Waist circumference, total cholesterol, triglyceride, low-density lipoprotein cholesterol, HOMA-IR were significantly higher and IPH Stumvoll index and ISI-composite were significantly lower in high-normal blood pressure group than in normal blood pressure group (P < 0.05). Systolic blood pressure (SBP) was positively correlated with HOMA-IR (r = 0.122) and negatively correlated with 1PH Stumvoll index (r = -0. 159) and ISI-composite (r = -0.131) (P < 0.05). SBP and triglyceride were independent factors for IPH Stumvoll index. Insulin resistance and islet dysfunction may exist in subjects with high-normal blood pressure.

Effects of Poloxamer 188 on red blood cell membrane properties in sickle cell anaemia.

PubMed

Sandor, Barbara; Marin, Mickaël; Lapoumeroulie, Claudine; Rabaï, Miklos; Lefevre, Sophie D; Lemonne, Nathalie; El Nemer, Wassim; Mozar, Anaïs; Français, Olivier; Le Pioufle, Bruno; Connes, Philippe; Le Van Kim, Caroline

2016-04-01

Vaso-occlusive crisis (VOC) is the main acute complication in sickle cell anaemia (SS) and several clinical trials are investigating different drugs to improve the clinical severity of SS patients. A phase III study is currently exploring the profit of Velopoloxamer in SS during VOCs. We analysed, in-vitro, the effect of poloxamer (P188) on red blood cell (RBC) properties by investigating haemorheology, mechanical and adhesion functions using ektacytometry, microfluidics and dynamic adhesion approaches, respectively. We show that poloxamer significantly reduces blood viscosity, RBC aggregation and adhesion to endothelial cells, supporting the beneficial use of this molecule in SS therapy. © 2016 John Wiley & Sons Ltd.

Biomarkers of splenic function in infants with sickle cell anemia: baseline data from the BABY HUG Trial.

PubMed

Rogers, Zora R; Wang, Winfred C; Luo, Zhaoyu; Iyer, Rathi V; Shalaby-Rana, Eglal; Dertinger, Stephen D; Shulkin, Barry L; Miller, John H; Files, Bea; Lane, Peter A; Thompson, Bruce W; Miller, Scott T; Ware, Russell E

2011-03-03

We evaluated spleen function in 193 children with sickle cell anemia 8 to 18 months of age by (99m)Tc sulfur-colloid liver-spleen scan and correlated results with clinical and laboratory parameters, including 2 splenic biomarkers: pitted cell counts (PIT) and quantitative Howell-Jolly bodies (HJB) enumerated by flow cytometry. Loss of splenic function began before 12 months of age in 86% of infants in association with lower total or fetal hemoglobin and higher white blood cell or reticulocyte counts, reinforcing the need for early diagnosis and diligent preventive care. PIT and HJB correlated well with each other and liver-spleen scan results. Previously described biomarker threshold values did define patients with abnormal splenic function, but our data suggest that normal spleen function is better predicted by PIT of ≤1.2% or HJB ≤55/10(6) red blood cells and absent function by PIT ≥4.5% or HJB ≥665/10(6). HJB is methodologically advantageous compared with PIT, but both are valid biomarkers of splenic function. This trial was registered at www.clinicaltrials.gov as #NCT00006400. © 2011 by The American Society of Hematology

Relationship between hyperglycemia, hormone disturbances, and clinical evolution in severely hyperglycemic post surgery critically ill children: an observational study

PubMed Central

2014-01-01

Background To study hormonal changes associated with severe hyperglycemia in critically ill children and the relationship with prognosis and length of stay in intensive care. Methods Observational study in twenty-nine critically ill children with severe hyperglycemia defined as 2 blood glucose measurements greater than 180 mg/dL. Severity of illness was assessed using pediatric index of mortality (PIM2), pediatric risk of mortality (PRISM) score, and pediatric logistic organ dysfunction (PELOD) scales. Blood glucose, glycosuria, insulin, C-peptide, cortisol, corticotropin, insulinlike growth factor-1, growth hormone, thyrotropin, thyroxine, and treatment with insulin were recorded. β-cell function and insulin sensitivity and resistance were determined on the basis of the homeostatic model assessment (HOMA), using blood glucose and C-peptide levels. Results The initial blood glucose level was 249 mg/dL and fell gradually to 125 mg/dL at 72 hours. Initial β-cell function (49.2%) and insulin sensitivity (13.2%) were low. At the time of diagnosis of hyperglycemia, 50% of the patients presented insulin resistance and β-cell dysfunction, 46% presented isolated insulin resistance, and 4% isolated β-cell dysfunction. β-cell function improved rapidly but insulin resistance persisted. Initial glycemia did not correlate with any other factor, and there was no relationship between glycemia and mortality. Patients who died had higher cortisol and growth hormone levels at diagnosis. Length of stay was correlated by univariate analysis, but not by multivariate analysis, with C-peptide and glycemic control at 24 hours, insulin resistance, and severity of illness scores. Conclusions Critically ill children with severe hyperglycemia initially present decreased β-cell function and insulin sensitivity. Nonsurvivors had higher cortisol and growth hormone levels and developed hyperglycemia later than survivors. PMID:24628829

Successful In Vitro Expansion and Differentiation of Cord Blood Derived CD34+ Cells into Early Endothelial Progenitor Cells Reveals Highly Differential Gene Expression

PubMed Central

Topcic, Denijal; Haviv, Izhak; Merivirta, Ruusu-Maaria; Agrotis, Alexander; Leitner, Ephraem; Jowett, Jeremy B.; Bode, Christoph; Lappas, Martha; Peter, Karlheinz

2011-01-01

Endothelial progenitor cells (EPCs) can be purified from peripheral blood, bone marrow or cord blood and are typically defined by a limited number of cell surface markers and a few functional tests. A detailed in vitro characterization is often restricted by the low cell numbers of circulating EPCs. Therefore in vitro culturing and expansion methods are applied, which allow at least distinguishing two different types of EPCs, early and late EPCs. Herein, we describe an in vitro culture technique with the aim to generate high numbers of phenotypically, functionally and genetically defined early EPCs from human cord blood. Characterization of EPCs was done by flow cytometry, immunofluorescence microscopy, colony forming unit (CFU) assay and endothelial tube formation assay. There was an average 48-fold increase in EPC numbers. EPCs expressed VEGFR-2, CD144, CD18, and CD61, and were positive for acetylated LDL uptake and ulex lectin binding. The cells stimulated endothelial tube formation only in co-cultures with mature endothelial cells and formed CFUs. Microarray analysis revealed highly up-regulated genes, including LL-37 (CAMP), PDK4, and alpha-2-macroglobulin. In addition, genes known to be associated with cardioprotective (GDF15) or pro-angiogenic (galectin-3) properties were also significantly up-regulated after a 72 h differentiation period on fibronectin. We present a novel method that allows to generate high numbers of phenotypically, functionally and genetically characterized early EPCs. Furthermore, we identified several genes newly linked to EPC differentiation, among them LL-37 (CAMP) was the most up-regulated gene. PMID:21858032

Hypergravity Alters the Susceptibility of Cells to Anoxia-Reoxygenation Injury

NASA Technical Reports Server (NTRS)

McCloud, Henry; Pink, Yulondo; Harris-Hooker, Sandra A.; Melhado, Caroline D.; Sanford, Gary L.

1997-01-01

Gravity is a physical force, much like shear stress or mechanical stretch, and should affect organ and cellular function. Researchers have shown that gravity plays a role in ventilation and blood flow distribution, gas exchange, alveolar size and mechanical stresses within the lung. Short exposure to microgravity produced marked alterations in lung blood flow and ventilation distribution while hypergravity exaggerated the regional differences in lung structure and function resulting in reduced ventilation at the base and no ventilation of the upper half of the lung. Microgravity also decreased metabolic activity in cardiac cells, WI-38 embryonic lung cells, and human lymphocytes. Rats, in the tail-suspended head-down tilt model, experienced transient loss of lung water, contrary to an expected increase due to pooling of blood in the pulmonary vasculature. Hypergravity has also been found to increase the proliferation of several different cell lines (e.g., chick embryo fibroblasts) while decreasing cell motility and slowing liver regeneration following partial hepatectomy. These studies show that changes in the gravity environment will affect several aspects of organ and cellular function and produce major change in blood flow and tissue/organ perfusion. However, these past studies have not addressed whether ischemia-reperfusion injury will be exacerbated or ameliorated by changes in the gravity environment, e.g., space flight. Currently, nothing is known about how gravity will affect the susceptibility of different lung and vascular cells to this type of injury. We conducted studies that addressed the following question: Does the susceptibility of lung fibroblasts, vascular smooth muscle, and endothelial cells to anoxia/reoxygenation injury change following exposure to hypergravity conditions?

Hypoxia modulates the purine salvage pathway and decreases red blood cell and supernatant levels of hypoxanthine during refrigerated storage.

PubMed

Nemkov, Travis; Sun, Kaiqi; Reisz, Julie A; Song, Anren; Yoshida, Tatsuro; Dunham, Andrew; Wither, Matthew J; Francis, Richard O; Roach, Robert C; Dzieciatkowska, Monika; Rogers, Stephen C; Doctor, Allan; Kriebardis, Anastasios; Antonelou, Marianna; Papassideri, Issidora; Young, Carolyn T; Thomas, Tiffany A; Hansen, Kirk C; Spitalnik, Steven L; Xia, Yang; Zimring, James C; Hod, Eldad A; D'Alessandro, Angelo

2018-02-01

Hypoxanthine catabolism in vivo is potentially dangerous as it fuels production of urate and, most importantly, hydrogen peroxide. However, it is unclear whether accumulation of intracellular and supernatant hypoxanthine in stored red blood cell units is clinically relevant for transfused recipients. Leukoreduced red blood cells from glucose-6-phosphate dehydrogenase-normal or -deficient human volunteers were stored in AS-3 under normoxic, hyperoxic, or hypoxic conditions (with oxygen saturation ranging from <3% to >95%). Red blood cells from healthy human volunteers were also collected at sea level or after 1-7 days at high altitude (>5000 m). Finally, C57BL/6J mouse red blood cells were incubated in vitro with 13 C 1 -aspartate or 13 C 5 -adenosine under normoxic or hypoxic conditions, with or without deoxycoformycin, a purine deaminase inhibitor. Metabolomics analyses were performed on human and mouse red blood cells stored for up to 42 or 14 days, respectively, and correlated with 24 h post-transfusion red blood cell recovery. Hypoxanthine increased in stored red blood cell units as a function of oxygen levels. Stored red blood cells from human glucose-6-phosphate dehydrogenase-deficient donors had higher levels of deaminated purines. Hypoxia in vitro and in vivo decreased purine oxidation and enhanced purine salvage reactions in human and mouse red blood cells, which was partly explained by decreased adenosine monophosphate deaminase activity. In addition, hypoxanthine levels negatively correlated with post-transfusion red blood cell recovery in mice and - preliminarily albeit significantly - in humans. In conclusion, hypoxanthine is an in vitro metabolic marker of the red blood cell storage lesion that negatively correlates with post-transfusion recovery in vivo Storage-dependent hypoxanthine accumulation is ameliorated by hypoxia-induced decreases in purine deamination reaction rates. Copyright© 2018 Ferrata Storti Foundation.

Hypoxia modulates the purine salvage pathway and decreases red blood cell and supernatant levels of hypoxanthine during refrigerated storage

PubMed Central

Nemkov, Travis; Sun, Kaiqi; Reisz, Julie A.; Song, Anren; Yoshida, Tatsuro; Dunham, Andrew; Wither, Matthew J.; Francis, Richard O.; Roach, Robert C.; Dzieciatkowska, Monika; Rogers, Stephen C.; Doctor, Allan; Kriebardis, Anastasios; Antonelou, Marianna; Papassideri, Issidora; Young, Carolyn T.; Thomas, Tiffany A.; Hansen, Kirk C.; Spitalnik, Steven L.; Xia, Yang; Zimring, James C.; Hod, Eldad A.; D’Alessandro, Angelo

2018-01-01

Hypoxanthine catabolism in vivo is potentially dangerous as it fuels production of urate and, most importantly, hydrogen peroxide. However, it is unclear whether accumulation of intracellular and supernatant hypoxanthine in stored red blood cell units is clinically relevant for transfused recipients. Leukoreduced red blood cells from glucose-6-phosphate dehydrogenase-normal or -deficient human volunteers were stored in AS-3 under normoxic, hyperoxic, or hypoxic conditions (with oxygen saturation ranging from <3% to >95%). Red blood cells from healthy human volunteers were also collected at sea level or after 1–7 days at high altitude (>5000 m). Finally, C57BL/6J mouse red blood cells were incubated in vitro with 13C1-aspartate or 13C5-adenosine under normoxic or hypoxic conditions, with or without deoxycoformycin, a purine deaminase inhibitor. Metabolomics analyses were performed on human and mouse red blood cells stored for up to 42 or 14 days, respectively, and correlated with 24 h post-transfusion red blood cell recovery. Hypoxanthine increased in stored red blood cell units as a function of oxygen levels. Stored red blood cells from human glucose-6-phosphate dehydrogenase-deficient donors had higher levels of deaminated purines. Hypoxia in vitro and in vivo decreased purine oxidation and enhanced purine salvage reactions in human and mouse red blood cells, which was partly explained by decreased adenosine monophosphate deaminase activity. In addition, hypoxanthine levels negatively correlated with post-transfusion red blood cell recovery in mice and – preliminarily albeit significantly - in humans. In conclusion, hypoxanthine is an in vitro metabolic marker of the red blood cell storage lesion that negatively correlates with post-transfusion recovery in vivo. Storage-dependent hypoxanthine accumulation is ameliorated by hypoxia-induced decreases in purine deamination reaction rates. PMID:29079593

Anomalies of the CD8+ T cell pool in haemochromatosis: HLA-A3-linked expansions of CD8+CD28- T cells.

PubMed

Arosa, F A; Oliveira, L; Porto, G; da Silva, B M; Kruijer, W; Veltman, J; de Sousa, M

1997-03-01

The present study consists of a phenotypic and functional characterization of peripheral blood T lymphocytes in a group of 21 patients with hereditary haemochromatosis (HH), an MHC class I-linked genetic disease resulting in iron overload, and a group of 30 healthy individuals, both HLA-phenotyped. The HH patients studied showed an increased percentage of CD8+ CD28- T cells with a corresponding reduction in the percentage of CD8+ CD28+ T cells in peripheral blood relative to healthy blood donors. No anomalies of CD28 expression were found in the CD4+ subset. The presence of the HLA-A3 antigen but not age accounted for these imbalances. Thus, an apparent failure of the CD8+ CD28+ T cell population 'to expand', coinciding with an 'expansion' of CD8+ CD28- T cells in peripheral blood of HLA-A3+ but not HLA-A3- HH patients was observed when compared with the respective HLA-A3-matched control group. A significantly higher percentage of HLA-DR+ but not CD45RO+ cells was also found within the peripheral CD8+ T cell subset in HH patients relative to controls. Phytohaemagglutinin (PHA) stimulation of peripheral blood mononuclear cells (PBMC) for 5 days showed: (i) that CD8+ CD28+ T cells both in controls and HH were able to expand in vitro; (ii) that CD8+ CD28- T cells decreased markedly after activation in controls but not in HH patients. Moreover, functional studies showed that CD8+ cytotoxic T lymphocytes (CTL) from HH patients exhibited a diminished cytotoxic activity (approx. two-fold) in standard 51Cr-release assays when compared with CD8+ CTL from healthy controls. The present results provide additional evidence for the existence of phenotypic and functional anomalies of the peripheral CD8+ T cell pool that may underlie the clinical heterogeneity of this iron overload disease. They are of particular relevance given the recent discovery of a novel mutated MHC class I-like gene in HH.

Anomalies of the CD8+ T cell pool in haemochromatosis: HLA-A3-linked expansions of CD8+CD28− T cells

PubMed Central

AROSA, F A; OLIVEIRA, L; PORTO, G; DA SILVA, B M; KRUIJER, W; VELTMAN, J; DE SOUSA, M

1997-01-01

The present study consists of a phenotypic and functional characterization of peripheral blood T lymphocytes in a group of 21 patients with hereditary haemochromatosis (HH), an MHC class I-linked genetic disease resulting in iron overload, and a group of 30 healthy individuals, both HLA-phenotyped. The HH patients studied showed an increased percentage of CD8+ CD28− T cells with a corresponding reduction in the percentage of CD8+ CD28+ T cells in peripheral blood relative to healthy blood donors. No anomalies of CD28 expression were found in the CD4+ subset. The presence of the HLA-A3 antigen but not age accounted for these imbalances. Thus, an apparent failure of the CD8+ CD28+ T cell population ‘to expand’, coinciding with an ‘expansion’ of CD8+ CD28− T cells in peripheral blood of HLA-A3+ but not HLA-A3− HH patients was observed when compared with the respective HLA-A3-matched control group. A significantly higher percentage of HLA-DR+ but not CD45RO+ cells was also found within the peripheral CD8+ T cell subset in HH patients relative to controls. Phytohaemagglutinin (PHA) stimulation of peripheral blood mononuclear cells (PBMC) for 5 days showed: (i) that CD8+ CD28+ T cells both in controls and HH were able to expand in vitro; (ii) that CD8+ CD28− T cells decreased markedly after activation in controls but not in HH patients. Moreover, functional studies showed that CD8+ cytotoxic T lymphocytes (CTL) from HH patients exhibited a diminished cytotoxic activity (approx. two-fold) in standard 51Cr-release assays when compared with CD8+ CTL from healthy controls. The present results provide additional evidence for the existence of phenotypic and functional anomalies of the peripheral CD8+ T cell pool that may underlie the clinical heterogeneity of this iron overload disease. They are of particular relevance given the recent discovery of a novel mutated MHC class I-like gene in HH. PMID:9067531

Cell-cell interaction in blood flow in patients with coronary heart disease (in vitro study)

NASA Astrophysics Data System (ADS)

Malinova, Lidia I.; Simonenko, Georgy V.; Denisova, Tatyana P.; Tuchin, Valery V.

2007-02-01

Blood cell-cell and cell-vessel wall interactions are one of the key patterns in blood and vascular pathophysiology. We have chosen the method of reconstruction of pulsative blood flow in vitro in the experimental set. Blood flow structure was studied by PC integrated video camera with following slide by slide analysis. Studied flow was of constant volumetric blood flow velocity (1 ml/h). Diameter of tube in use was comparable with coronary arteries diameter. Glucose solution and unfractured heparin were used as the nonspecial irritants of studied flow. Erythrocytes space structure in flow differs in all groups of patients in our study (men with stable angina pectoris (SAP), myocardial infarction (MI) and practically healthy men (PHM). Intensity of erythrocytes aggregate formation was maximal in patients with SAP, but time of their "construction/deconstruction" at glucose injection was minimal. Phenomena of primary clotting formation in patients with SAP of high function class was reconstructed under experimental conditions. Heparin injection (10 000 ED) increased linear blood flow velocity both in patients with SAP, MI and PHP but modulated the cell profile in the flow. Received data correspond with results of animal model studies and noninvasive blood flow studies in human. Results of our study reveal differences in blood flow structure in patients with coronary heart disease and PHP under irritating conditions as the possible framework of metabolic model of coronary blood flow destabilization.

Relative deformability of red blood cells in sickle cell trait and sickle cell anemia by trapping and dragging

NASA Astrophysics Data System (ADS)

Solomon, Rance; Cooper, James; Welker, Gabriel; Aguilar, Elaura; Flanagan, Brooke; Pennycuff, Chelsey; Scott, David; Farone, Anthony; Farone, Mary; Erenso, Daniel; Mushi, Robert; del Pilar Aguinaga, Maria

2013-06-01

Genetic mutation of the β-globin gene or inheritance of this mutated gene changes the chemical composition of the oxygen-carrying hemoglobin molecule that could lead to either the heterozygote genotype, resulting in sickle cell trait (SCT), or the homozygote genotype, resulting in sickle cell anemia (SCA). These mutations could affect the reversible elastic deformations of the red blood cells (RBCs) which are vital for biological functions. We have investigated this effect by studying the differences in the deformability of RBCs from blood samples of an individual with SCT and an untreated patient with SCA along with hemoglobin quantitation of each blood sample. Infrared 1064 nm laser trap force along with drag shear force are used to induce deformation in the RBCs. Ultra2-High Performance Liquid Chromatography (UHPLC) is used for the hemoglobin quantitation.

[Whole-blood transfusion for hemorrhagic shock resuscitation: two cases in Djibouti].

PubMed

Cordier, P Y; Eve, O; Dehan, C; Topin, F; Menguy, P; Bertani, A; Massoure, P L; Kaiser, E

2012-01-01

Hemorrhagic shock requires early aggressive treatment, including transfusion of packed red blood cells and hemostatic resuscitation. In austere environments, when component therapy is not available, warm fresh whole-blood transfusion is a convenient treatment. It provides red blood cells, clotting factors, and functional platelets. Therefore it is commonly used in military practice to treat hemorrhagic shock in combat casualties. At Bouffard Hospital Center in Djibouti, the supply of packed red blood cells is limited, and apheresis platelets are unavailable. We used whole blood transfusion in two civilian patients with life-threatening non-traumatic hemorrhages. One had massive bleeding caused by disseminated intravascular coagulation due to septic shock; the second was a 39 year-old pregnant woman with uterine rupture. In both cases, whole blood transfusion (twelve and ten 500 mL bags respectively), combined with etiological treatment, enabled coagulopathy correction, hemorrhage control, and satisfactory recovery.

High Endothelial Venules and Other Blood Vessels: Critical Regulators of Lymphoid Organ Development and Function

PubMed Central

Ager, Ann

2017-01-01

The blood vasculature regulates both the development and function of secondary lymphoid organs by providing a portal for entry of hemopoietic cells. During the development of lymphoid organs in the embryo, blood vessels deliver lymphoid tissue inducer cells that initiate and sustain the development of lymphoid tissues. In adults, the blood vessels are structurally distinct from those in other organs due to the requirement for high levels of lymphocyte recruitment under non-inflammatory conditions. In lymph nodes (LNs) and Peyer’s patches, high endothelial venules (HEVs) especially adapted for lymphocyte trafficking form a spatially organized network of blood vessels, which controls both the type of lymphocyte and the site of entry into lymphoid tissues. Uniquely, HEVs express vascular addressins that regulate lymphocyte entry into lymphoid organs and are, therefore, critical to the function of lymphoid organs. Recent studies have demonstrated important roles for CD11c+ dendritic cells in the induction, as well as the maintenance, of vascular addressin expression and, therefore, the function of HEVs. Tertiary lymphoid organs (TLOs) are HEV containing LN-like structures that develop inside organized tissues undergoing chronic immune-mediated inflammation. In autoimmune lesions, the development of TLOs is thought to exacerbate disease. In cancerous tissues, the development of HEVs and TLOs is associated with improved patient outcomes in several cancers. Therefore, it is important to understand what drives the development of HEVs and TLOs and how these structures contribute to pathology. In several human diseases and experimental animal models of chronic inflammation, there are some similarities between the development and function of HEVs within LN and TLOs. This review will summarize current knowledge of how hemopoietic cells with lymphoid tissue-inducing, HEV-inducing, and HEV-maintaining properties are recruited from the bloodstream to induce the development and control the function of lymphoid organs. PMID:28217126

[Clinical effect of stem cell transplantation via hepatic artery in the treatment of type II hyperammonemia: a report on 6 cases].

PubMed

DU, Kan; Luan, Zuo; Qu, Su-Qing; Yang, Hui; Yang, Yin-Xiang; Wang, Zhao-Yan; Jin, Hui-Yu; Liu, Wei-Peng

2013-11-01

This study aimed to investigate the clinical effect of transplantation of CD133⁺ peripheral blood stem cells or umbilical cord mesenchymal stem cells via the hepatic artery in children with type II hyperammonemia and its possible action mechanism. Umbilical cord mesenchymal stem cells were obtained by collecting cord blood (100-150 mL) from healthy fetuses and separating stem cell suspension (5 mL) from the cord blood by hydroxyethyl starch sedimentation. CD133⁺ peripheral blood stem cells were obtained by mobilizing peripheral blood from the fathers of sick children using recombinant human granulocyte colony-stimulating factor for 5 days, collecting mononuclear cells (120 mL), and separating out CD133⁺ cells by sorting. With catheterization and percutaneous puncture, the obtained stem cells were slowly injected into the liver of sick children via the hepatic artery. The changes in clinical symptoms and laboratory indices such as blood ammonia, liver function, and arginine and citrulline concentrations were observed. After stem cell transplantation via the hepatic artery, the 6 children showed significantly decreased blood ammonia levels, and their blood ammonia levels slowly increased 1 to 2 weeks later, but remained below 100 μmol/L, and changes in glutamic-pyruvic transaminase levels were similar to blood ammonia. Plasma citrulline and arginine concentrations increased significantly after transplantation and the increase in citrulline level exceeded the increase in arginine level. An 8 months follow-up visit for one typical patient showed that the weight and height increased after transplantation and sleep was improved without night crying. The child could actively gaze at interesting objects instead of responding indifferently and started to say simple words. With regard to fine motor skills, the child could pinch things with the thumb and middle finger instead of displaying a lack of hand-eye coordination and progress was also made in gross motor skills. Gesell test showed that the child made progress for an average of 3.82 months in all areas. It was concluded that after stem cell transplantation, children with type II hyperammonemia have decreased blood ammonia levels, stable and improved liver function and steadily increased plasma citrulline and arginine concentrations. They display a progressive trend in such aspects as movement, language and environmental adaptability. It is hypothesized that stem cell transplantation via the hepatic artery partially or totally activates, or provides supplementary ornithine carbamoyl transferase, so that plasma citrulline and arginine concentrations increase and urea cycle disorder can be corrected to some extent.

Metabolism of Citrate and Other Carboxylic Acids in Erythrocytes As a Function of Oxygen Saturation and Refrigerated Storage.

PubMed

Nemkov, Travis; Sun, Kaiqi; Reisz, Julie A; Yoshida, Tatsuro; Dunham, Andrew; Wen, Edward Y; Wen, Alexander Q; Roach, Rob C; Hansen, Kirk C; Xia, Yang; D'Alessandro, Angelo

2017-01-01

State-of-the-art proteomics technologies have recently helped to elucidate the unanticipated complexity of red blood cell metabolism. One recent example is citrate metabolism, which is catalyzed by cytosolic isoforms of Krebs cycle enzymes that are present and active in mature erythrocytes and was determined using quantitative metabolic flux analysis. In previous studies, we reported significant increases in glycolytic fluxes in red blood cells exposed to hypoxia in vitro or in vivo , an observation relevant to transfusion medicine owing to the potential benefits associated with hypoxic storage of packed red blood cells. Here, using a combination of steady state and quantitative tracing metabolomics experiments with 13 C 1,2,3 -glucose, 13 C 6 -citrate, 13 C 5 15 N 2 -glutamine, and 13 C 1 -aspartate via ultra-high performance liquid chromatography coupled on line with mass spectrometry, we observed that hypoxia in vivo and in vitro promotes consumption of citrate and other carboxylates. These metabolic reactions are theoretically explained by the activity of cytosolic malate dehydrogenase 1 and isocitrate dehydrogenase 1 (abundantly represented in the red blood cell proteome), though moonlighting functions of additional enzymes cannot be ruled out. These observations enhance understanding of red blood cell metabolic responses to hypoxia, which could be relevant to understand systemic physiological and pathological responses to high altitude, ischemia, hemorrhage, sepsis, pulmonary hypertension, or hemoglobinopathies. Results from this study will also inform the design and testing of novel additive solutions that optimize red blood cell storage under oxygen-controlled conditions.

Metabolism of Citrate and Other Carboxylic Acids in Erythrocytes As a Function of Oxygen Saturation and Refrigerated Storage

PubMed Central

Nemkov, Travis; Sun, Kaiqi; Reisz, Julie A.; Yoshida, Tatsuro; Dunham, Andrew; Wen, Edward Y.; Wen, Alexander Q.; Roach, Rob C.; Hansen, Kirk C.; Xia, Yang; D’Alessandro, Angelo

2017-01-01

State-of-the-art proteomics technologies have recently helped to elucidate the unanticipated complexity of red blood cell metabolism. One recent example is citrate metabolism, which is catalyzed by cytosolic isoforms of Krebs cycle enzymes that are present and active in mature erythrocytes and was determined using quantitative metabolic flux analysis. In previous studies, we reported significant increases in glycolytic fluxes in red blood cells exposed to hypoxia in vitro or in vivo, an observation relevant to transfusion medicine owing to the potential benefits associated with hypoxic storage of packed red blood cells. Here, using a combination of steady state and quantitative tracing metabolomics experiments with 13C1,2,3-glucose, 13C6-citrate, 13C515N2-glutamine, and 13C1-aspartate via ultra-high performance liquid chromatography coupled on line with mass spectrometry, we observed that hypoxia in vivo and in vitro promotes consumption of citrate and other carboxylates. These metabolic reactions are theoretically explained by the activity of cytosolic malate dehydrogenase 1 and isocitrate dehydrogenase 1 (abundantly represented in the red blood cell proteome), though moonlighting functions of additional enzymes cannot be ruled out. These observations enhance understanding of red blood cell metabolic responses to hypoxia, which could be relevant to understand systemic physiological and pathological responses to high altitude, ischemia, hemorrhage, sepsis, pulmonary hypertension, or hemoglobinopathies. Results from this study will also inform the design and testing of novel additive solutions that optimize red blood cell storage under oxygen-controlled conditions. PMID:29090212

Mitochondrial dysfunction in blood cells from amyotrophic lateral sclerosis patients.

PubMed

Ehinger, Johannes K; Morota, Saori; Hansson, Magnus J; Paul, Gesine; Elmér, Eskil

2015-06-01

Mitochondrial dysfunction is implicated in amyotrophic lateral sclerosis, where the progressive degeneration of motor neurons results in muscle atrophy, paralysis and death. Abnormalities in both central nervous system and muscle mitochondria have previously been demonstrated in patient samples, indicating systemic disease. In this case-control study, venous blood samples were acquired from 24 amyotrophic lateral sclerosis patients and 21 age-matched controls. Platelets and peripheral blood mononuclear cells were isolated and mitochondrial oxygen consumption measured in intact and permeabilized cells with additions of mitochondrial substrates, inhibitors and titration of an uncoupler. Respiratory values were normalized to cell count and for two markers of cellular mitochondrial content, citrate synthase activity and mitochondrial DNA, respectively. Mitochondrial function was correlated with clinical staging of disease severity. Complex IV (cytochrome c-oxidase)-activity normalized to mitochondrial content was decreased in platelets from amyotrophic lateral sclerosis patients both when normalized to citrate synthase activity and mitochondrial DNA copy number. In mononuclear cells, complex IV-activity was decreased when normalized to citrate synthase activity. Mitochondrial content was increased in amyotrophic lateral sclerosis patient platelets. In mononuclear cells, complex I activity declined and mitochondrial content increased progressively with advancing disease stage. The findings are, however, based on small subsets of patients and need to be confirmed. We conclude that when normalized to mitochondria-specific content, complex IV-activity is reduced in blood cells from amyotrophic lateral sclerosis patients and that there is an apparent compensatory increase in cellular mitochondrial content. This supports systemic involvement in amyotrophic lateral sclerosis and suggests further study of mitochondrial function in blood cells as a future biomarker for the disease.

Pancreas-Specific Sirt1-Deficiency in Mice Compromises Beta-Cell Function without Development of Hyperglycemia.

PubMed

Pinho, Andreia V; Bensellam, Mohammed; Wauters, Elke; Rees, Maxine; Giry-Laterriere, Marc; Mawson, Amanda; Ly, Le Quan; Biankin, Andrew V; Wu, Jianmin; Laybutt, D Ross; Rooman, Ilse

2015-01-01

Sirtuin 1 (Sirt1) has been reported to be a critical positive regulator of glucose-stimulated insulin secretion in pancreatic beta-cells. The effects on islet cells and blood glucose levels when Sirt1 is deleted specifically in the pancreas are still unclear. This study examined islet glucose responsiveness, blood glucose levels, pancreatic islet histology and gene expression in Pdx1Cre; Sirt1ex4F/F mice that have loss of function and loss of expression of Sirt1 specifically in the pancreas. We found that in the Pdx1Cre; Sirt1ex4F/F mice, the relative insulin positive area and the islet size distribution were unchanged. However, beta-cells were functionally impaired, presenting with lower glucose-stimulated insulin secretion. This defect was not due to a reduced expression of insulin but was associated with a decreased expression of the glucose transporter Slc2a2/Glut2 and of the Glucagon like peptide-1 receptor (Glp1r) as well as a marked down regulation of endoplasmic reticulum (ER) chaperones that participate in the Unfolded Protein Response (UPR) pathway. Counter intuitively, the Sirt1-deficient mice did not develop hyperglycemia. Pancreatic polypeptide (PP) cells were the only other islet cells affected, with reduced numbers in the Sirt1-deficient pancreas. This study provides new mechanistic insights showing that beta-cell function in Sirt1-deficient pancreas is affected due to altered glucose sensing and deregulation of the UPR pathway. Interestingly, we uncovered a context in which impaired beta-cell function is not accompanied by increased glycemia. This points to a unique compensatory mechanism. Given the reduction in PP, investigation of its role in the control of blood glucose is warranted.

An optimized protocol for the generation and functional analysis of human mast cells from CD34+ enriched cell populations.

PubMed

Yin, Yuzhi; Bai, Yun; Olivera, Ana; Desai, Avanti; Metcalfe, Dean D

2017-09-01

The culture of mast cells from human tissues such a cord blood, peripheral blood or bone marrow aspirates has advanced our understanding of human mast cells (huMC) degranulation, mediator production and response to pharmacologic agents. However, existing methods for huMC culture tend to be laborious and expensive. Combining technical approaches from several of these protocols, we designed a simplified and more cost effective approach to the culture of mast cells from human cell populations including peripheral blood and cryopreserved cells from lymphocytapheresis. On average, we reduced by 30-50 fold the amount of culture media compared to our previously reported method, while the total MC number generated by this method (2.46±0.63×10 6 vs. 2.4±0.28×10 6 , respectively, from 1.0×10 8 lymphocytapheresis or peripheral blood mononuclear blood cells [PBMCs]) was similar to our previous method (2.36±0.70×10 6 ), resulting in significant budgetary savings. In addition, we compared the yield of huMCs with or without IL-3 added to early cultures in the presence of stem cell factor (SCF) and interlukin-6 (IL-6) and found that the total MC number generated, while higher with IL-3 in the culture, did not reach statistical significance, suggesting that IL-3, often recommended in the culture of huMCs, is not absolutely required. We then performed a functional analysis by flow cytometry using standard methods and which maximized the data we could obtain from cultured cells. We believe these approaches will allow more laboratories to culture and examine huMC behavior going forward. Published by Elsevier B.V.

Mucosa-associated invariant T cells link intestinal immunity with antibacterial immune defects in alcoholic liver disease.

PubMed

Riva, Antonio; Patel, Vishal; Kurioka, Ayako; Jeffery, Hannah C; Wright, Gavin; Tarff, Sarah; Shawcross, Debbie; Ryan, Jennifer M; Evans, Alexander; Azarian, Sarah; Bajaj, Jasmohan S; Fagan, Andrew; Patel, Vinood; Mehta, Kosha; Lopez, Carlos; Simonova, Marieta; Katzarov, Krum; Hadzhiolova, Tanya; Pavlova, Slava; Wendon, Julia A; Oo, Ye Htun; Klenerman, Paul; Williams, Roger; Chokshi, Shilpa

2018-05-01

Intestinal permeability with systemic distribution of bacterial products are central in the immunopathogenesis of alcoholic liver disease (ALD), yet links with intestinal immunity remain elusive. Mucosa-associated invariant T cells (MAIT) are found in liver, blood and intestinal mucosa and are a key component of antibacterial host defences. Their role in ALD is unknown. We analysed frequency, phenotype, transcriptional regulation and function of blood MAIT cells in severe alcoholic hepatitis (SAH), alcohol-related cirrhosis (ARC) and healthy controls (HC). We also examined direct impact of ethanol, bacterial products from faecal extracts and antigenic hyperstimulation on MAIT cell functionality. Presence of MAIT cells in colon and liver was assessed by quantitative PCR and immunohistochemistry/gene expression respectively. In ARC and SAH, blood MAIT cells were dramatically depleted, hyperactivated and displayed defective antibacterial cytokine/cytotoxic responses. These correlated with suppression of lineage-specific transcription factors and hyperexpression of homing receptors in the liver with intrahepatic preservation of MAIT cells in ALD. These alterations were stronger in SAH, where surrogate markers of bacterial infection and microbial translocation were higher than ARC. Ethanol exposure in vitro, in vivo alcohol withdrawal and treatment with Escherichia coli had no effect on MAIT cell frequencies, whereas exposure to faecal bacteria/antigens induced functional impairments comparable with blood MAIT cells from ALD and significant MAIT cell depletion, which was not observed in other T cell compartments. In ALD, the antibacterial potency of MAIT cells is compromised as a consequence of contact with microbial products and microbiota, suggesting that the 'leaky' gut observed in ALD drives MAIT cell dysfunction and susceptibility to infection in these patients. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Space-time Considerations for Hematopoietic Stem Cell Transplantation

PubMed Central

Bhattacharya, Deepta; Ehrlich, Lauren I. Richie; Weissman, Irving L.

2009-01-01

Summary The mammalian blood system contains a multitude of distinct mature cell lineages adapted to serve diverse functional roles. Mutations that abrogate the development or function of one or more of these lineages can lead to profound adverse consequences, such as immunodeficiency, autoimmunity, or anemia. Replacement of hematopoietic stem cells (HSCs) that carry such mutations with HSCs from a healthy donor can reverse such disorders, but because the risks associated with the procedure are often more serious than the blood disorders themselves, bone marrow transplantation is generally not used to treat a number of relatively common inherited blood diseases. Aside from a number of other problems, risks associated with cytoreductive treatments that create "space" for donor HSCs, and the slow kinetics with which immune competence is restored following transplantation hamper progress. This review will focus on how recent studies using experimental model systems may direct future efforts to implement routine use of HSC transplantation to cure inherited blood disorders. PMID:18651698

Human liver infiltrating γδ T cells are composed of clonally expanded circulating and tissue-resident populations.

PubMed

Hunter, Stuart; Willcox, Carrie R; Davey, Martin S; Kasatskaya, Sofya A; Jeffery, Hannah C; Chudakov, Dmitriy M; Oo, Ye H; Willcox, Benjamin E

2018-05-18

γδ T-cells comprise a substantial proportion of tissue-associated lymphocytes. However, our current understanding of human γδ T-cells is primarily based on peripheral blood subsets, while the immunobiology of tissue-associated subsets remains largely unclear. To address this, we characterised the TCR diversity, immunophenotype and function of human liver infiltrating γδ T-cells, focussing on the predominant tissue-associated Vδ2 neg γδ subset, which is implicated in liver immunopathology. Intrahepatic Vδ2 neg γδ T-cells were highly clonally focussed, with single expanded clonotypes featuring complex, private TCR rearrangements frequently dominating the compartment. Such T-cells were predominantly CD27 lo/neg effector lymphocytes, whereas naïve CD27 hi , TCR diverse populations present in matched blood were generally absent in the liver. Furthermore, while a CD45RA hi Vδ2 neg γδ effector subset present in both liver and peripheral blood contained overlapping TCR clonotypes, the liver Vδ2 neg γδ T-cell pool also included a phenotypically distinct CD45RA lo effector compartment that was enriched for expression of the tissue tropism marker CD69, the hepatic homing chemokine receptors CXCR3 and CXCR6, and liver-restricted TCR clonotypes, suggestive of intrahepatic tissue residency. Liver infiltrating Vδ2 neg γδ cells were capable of polyfunctional cytokine secretion, and unlike peripheral blood subsets, were responsive to both TCR and innate stimuli. These findings suggest the ability of Vδ2 neg γδ T-cells to undergo clonotypic expansion and differentiation is crucial in permitting access to solid tissues such as the liver, and can result in functionally distinct peripheral and liver-resident memory γδ T-cell subsets. They highlight the inherent functional plasticity within the Vδ2 neg γδ T-cell compartment, and may inform design of cellular therapies involving intrahepatic trafficking of γδ T-cells to suppress liver inflammation or combat liver cancer. γδ T cells are frequently enriched in many solid tissues, however the immunobiology of such tissue-associated subsets in humans has remained unclear. We show that intrahepatic γδ T cells are enriched for clonally expanded effector T cells, whereas naïve γδ T cells are largely excluded; moreover, whereas a distinct proportion of circulating T cell clonotypes was present in both the liver tissue and peripheral blood, a functionally and clonotypically distinct population of liver-resident γδ T cells was also evident. Our findings suggest that factors triggering γδ T cell clonal selection and differentiation, such as infection, can drive enrichment of γδ T cells into liver tissue, allowing the development of functionally distinct tissue-restricted memory populations specialised in local hepatic immunosurveillance. Copyright © 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Transcriptional regulation of pancreas development and β-cell function [Review].

PubMed

Fujitani, Yoshio

2017-05-30

A small number of cells in the adult pancreas are endocrine cells. They are arranged in clusters called islets of Langerhans. The islets make insulin, glucagon, and other endocrine hormones, and release them into the blood circulation. These hormones help control the level of blood glucose. Therefore, a dysfunction of endocrine cells in the pancreas results in impaired glucose homeostasis, or diabetes mellitus. The pancreas is an organ that originates from the evaginations of pancreatic progenitor cells in the epithelium of the foregut endoderm. Pancreas organogenesis and maturation of the islets of Langerhans occurs via a coordinated and complex interplay of transcriptional networks and signaling molecules, which guide a stepwise and repetitive process of the propagation of progenitor cells and their maturation, eventually resulting in a fully functional organ. Increasing our understanding of the extrinsic, as well as intrinsic mechanisms that control these processes should facilitate the efforts to generate surrogate β cells from ES or iPS cells, or to reactivate the function of important cell types within pancreatic islets that are lost in diabetes.

Genetics Home Reference: retroperitoneal fibrosis

MedlinePlus

... substances build up in the blood and tissues, leading to nausea, vomiting, weight loss, itching, a low number of red blood cells ( anemia ), and changes in brain function. Related Information What does it ...

Engineering cord blood to improve engraftment after cord blood transplant

PubMed Central

Dave, Hema; Bollard, Catherine M.; Shpall, Elizabeth J.

2017-01-01

Umbilical cord blood transplant (CBT) has traditionally been associated with slower engraftment of neutrophils, delayed immune reconstitution and consequently higher risk of infections as compared with peripheral blood progenitor cell (PBPC) or bone marrow (BM) transplants. This is primarily due to low numbers of total nucleated cells (TNCs) and the naive nature of CB immune cells. The use of double unit CB transplant (DCBT) increases the total cell dose in the graft, but it still does not produce as rapid engraftment as seen with PBPC or even BM transplants. Herein, we discuss strategies to improve engraftment after CBT. We describe methods of (I) expansion of CB graft ex vivo to increase the total cell dose; and (II) enhancement of BM homing capability of CB progenitor cells; (III) ex vivo expansion of CB derived T cells for improving T cell function against viruses, tumors and protection from graft versus host disease (GVHD). With these novel approaches, engraftment after CBT is now reaching levels comparable to that of other graft types. PMID:28607915

Quantitative investigation of red blood cell three-dimensional geometric and chemical changes in the storage lesion using digital holographic microscopy.

PubMed

Jaferzadeh, Keyvan; Moon, Inkyu

2015-11-01

Quantitative phase information obtained by digital holographic microscopy (DHM) can provide new insight into the functions and morphology of single red blood cells (RBCs). Since the functionality of a RBC is related to its three-dimensional (3-D) shape, quantitative 3-D geometric changes induced by storage time can help hematologists realize its optimal functionality period. We quantitatively investigate RBC 3-D geometric changes in the storage lesion using DHM. Our experimental results show that the substantial geometric transformation of the biconcave-shaped RBCs to the spherocyte occurs due to RBC storage lesion. This transformation leads to progressive loss of cell surface area, surface-to-volume ratio, and functionality of RBCs. Furthermore, our quantitative analysis shows that there are significant correlations between chemical and morphological properties of RBCs.

Clinical Response of 277 Patients with Spinal Cord Injury to Stem Cell Therapy in Iraq

PubMed Central

Hammadi, Abdulmajeed Alwan; Marino, Andolina; Farhan, Saad

2012-01-01

Background and Objectives: Spinal cord injury is a common neurological problem secondary to car accidents, war injuries and other causes, it may lead to varying degrees of neurological disablement, and apart from physiotherapy there is no available treatment to regain neurological function loss. Our aim is to find a new method using autologous hematopoietic stem cells to gain some of the neurologic functions lost after spinal cord injury. Methods and Results: 277 patients suffering from spinal cord injury were submitted to an intrathecally treatment with peripheral stem cells. The cells were harvested from the peripheral blood after a treatment with G-CSF and then concentrated to 4∼ 6 ml. 43% of the patients improved; ASIA score shifted from A to B in 88 and from A to C in 32. The best results were achieved in patients treated within one year from the injury. Conclusions: Since mesenchymal cells increase in the peripheral blood after G-CSF stimulation, a peripheral blood harvest seems easier and cheaper than mesenchymal cell cultivation prior to injection. It seems reasonable treatment for spinal cord injury. PMID:24298358

Pathogenic prion protein fragment (PrP106-126) promotes human immunodeficiency virus type-1 infection in peripheral blood monocyte-derived macrophages.

PubMed

Bacot, Silvia M; Feldman, Gerald M; Yamada, Kenneth M; Dhawan, Subhash

2015-02-01

Transfusion of blood and blood products contaminated with the pathogenic form of prion protein Prp(sc), thought to be the causative agent of variant a Creutzfeldt-Jakob disease (vCJD), may result in serious consequences in recipients with a compromised immune system, for example, as seen in HIV-1 infection. In the present study, we demonstrate that treatment of peripheral blood monocyte-derived macrophages (MDM) with PrP106-126, a synthetic domain of PrP(sc) that has intrinsic functional activities related to the full-length protein, markedly increased their susceptibility to HIV-1 infection, induced cytokine secretion, and enhanced their migratory behavior in response to N-formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP). Live-cell imaging of MDM cultured in the presence of PrP106-126 showed large cell clusters indicative of cellular activation. Tyrosine kinase inhibitor STI-571, protein kinase C inhibitor K252B, and cyclin-dependent kinase inhibitor olomoucine attenuated PrP106-126-induced altered MDM functions. These findings delineate a previously undefined functional role of PrP106-126-mediated host cell response in promoting HIV-1 pathogenesis. Published by Elsevier Inc.

Transplantation of in vitro cultured endothelial progenitor cells repairs the blood-brain barrier and improves cognitive function of APP/PS1 transgenic AD mice.

PubMed

Zhang, Shishuang; Zhi, Yongle; Li, Fei; Huang, Shan; Gao, Huabin; Han, Zhaoli; Ge, Xintong; Li, Dai; Chen, Fanglian; Kong, Xiaodong; Lei, Ping

2018-04-15

To date, the pathogenesis of Alzheimer's disease (AD) remains unclear. It is well-known that excessive deposition of Aβ in the brain is a crucial part of the pathogenesis of AD. In recent years, the AD neurovascular unit hypothesis has attracted much attention. Impairment of the blood-brain barrier (BBB) leads to abnormal amyloid-β (Aβ) transport, and chronic cerebral hypoperfusion causes Aβ deposition throughout the onset and progression of AD. Endothelial progenitor cells (EPCs) are the universal cells for repairing blood vessels. Our previous studies have shown that a reduced number of EPCs in the peripheral blood results in cerebral vascular repair disorder, cerebral hypoperfusion and neurodegeneration, which might be related to the cognitive dysfunction of AD patients. This study was designed to confirm whether EPCs transplantation could repair the blood-brain barrier, stimulate angiogenesis and reduce Aβ deposition in AD. The expression of ZO-1, Occludin and Claudin-5 was up-regulated in APP/PS1 transgenic mice after hippocampal transplantation of EPCs. Consistent with previous studies, EPC transplants also increased the microvessel density. We observed that Aβ senile plaque deposition was decreased and hippocampal cell apoptosis was reduced after EPCs transplantation. The Morris water maze test showed that spatial learning and memory functions were significantly improved in mice transplanted with EPCs. Consequently, EPCs could up-regulate the expression of tight junction proteins, repair BBB tight junction function, stimulate angiogenesis, promote Aβ clearance, and decrease neuronal loss, ultimately improve cognitive function. Taken together, these data demonstrate EPCs may play an important role in the therapeutic implications for vascular dysfunction in AD. Copyright © 2018 Elsevier B.V. All rights reserved.

Sourcing of an alternative pericyte-like cell type from peripheral blood in clinically relevant numbers for therapeutic angiogenic applications.

PubMed

Blocki, Anna; Wang, Yingting; Koch, Maria; Goralczyk, Anna; Beyer, Sebastian; Agarwal, Nikita; Lee, Michelle; Moonshi, Shehzahdi; Dewavrin, Jean-Yves; Peh, Priscilla; Schwarz, Herbert; Bhakoo, Kishore; Raghunath, Michael

2015-03-01

Autologous cells hold great potential for personalized cell therapy, reducing immunological and risk of infections. However, low cell counts at harvest with subsequently long expansion times with associated cell function loss currently impede the advancement of autologous cell therapy approaches. Here, we aimed to source clinically relevant numbers of proangiogenic cells from an easy accessible cell source, namely peripheral blood. Using macromolecular crowding (MMC) as a biotechnological platform, we derived a novel cell type from peripheral blood that is generated within 5 days in large numbers (10-40 million cells per 100 ml of blood). This blood-derived angiogenic cell (BDAC) type is of monocytic origin, but exhibits pericyte markers PDGFR-β and NG2 and demonstrates strong angiogenic activity, hitherto ascribed only to MSC-like pericytes. Our findings suggest that BDACs represent an alternative pericyte-like cell population of hematopoietic origin that is involved in promoting early stages of microvasculature formation. As a proof of principle of BDAC efficacy in an ischemic disease model, BDAC injection rescued affected tissues in a murine hind limb ischemia model by accelerating and enhancing revascularization. Derived from a renewable tissue that is easy to collect, BDACs overcome current short-comings of autologous cell therapy, in particular for tissue repair strategies.

Sourcing of an Alternative Pericyte-Like Cell Type from Peripheral Blood in Clinically Relevant Numbers for Therapeutic Angiogenic Applications

PubMed Central

Blocki, Anna; Wang, Yingting; Koch, Maria; Goralczyk, Anna; Beyer, Sebastian; Agarwal, Nikita; Lee, Michelle; Moonshi, Shehzahdi; Dewavrin, Jean-Yves; Peh, Priscilla; Schwarz, Herbert; Bhakoo, Kishore; Raghunath, Michael

2015-01-01

Autologous cells hold great potential for personalized cell therapy, reducing immunological and risk of infections. However, low cell counts at harvest with subsequently long expansion times with associated cell function loss currently impede the advancement of autologous cell therapy approaches. Here, we aimed to source clinically relevant numbers of proangiogenic cells from an easy accessible cell source, namely peripheral blood. Using macromolecular crowding (MMC) as a biotechnological platform, we derived a novel cell type from peripheral blood that is generated within 5 days in large numbers (10–40 million cells per 100 ml of blood). This blood-derived angiogenic cell (BDAC) type is of monocytic origin, but exhibits pericyte markers PDGFR-β and NG2 and demonstrates strong angiogenic activity, hitherto ascribed only to MSC-like pericytes. Our findings suggest that BDACs represent an alternative pericyte-like cell population of hematopoietic origin that is involved in promoting early stages of microvasculature formation. As a proof of principle of BDAC efficacy in an ischemic disease model, BDAC injection rescued affected tissues in a murine hind limb ischemia model by accelerating and enhancing revascularization. Derived from a renewable tissue that is easy to collect, BDACs overcome current short-comings of autologous cell therapy, in particular for tissue repair strategies. PMID:25582709

Patient-specific cardiovascular progenitor cells derived from integration-free induced pluripotent stem cells for vascular tissue regeneration.

PubMed

Hu, Jiang; Wang, Yongyu; Jiao, Jiao; Liu, Zhongning; Zhao, Chao; Zhou, Zhou; Zhang, Zhanpeng; Forde, Kaitlynn; Wang, Lunchang; Wang, Jiangang; Baylink, David J; Zhang, Xiao-Bing; Gao, Shaorong; Yang, Bo; Chen, Y Eugene; Ma, Peter X

2015-12-01

Tissue-engineered blood vessels (TEBVs) are promising in regenerating a live vascular replacement. However, the vascular cell source is limited, and it is crucial to develop a scaffold that accommodates new type of vascular progenitor cells and facilitates in vivo lineage specification of the cells into functional vascular smooth muscle cells (VSMCs) to regenerate vascular tissue. In the present study, integration-free human induced pluripotent stem cells (hiPSCs) were established from patient peripheral blood mononuclear cells through episomal vector nucleofection of reprogramming factors. The established hiPSCs were then induced into mesoderm-originated cardiovascular progenitor cells (CVPCs) with a highly efficient directed lineage specification method. The derived CVPCs were demonstrated to be able to differentiate into functional VSMCs. Subcutaneous implantation of CVPCs seeded on macroporous nanofibrous poly(l-lactide) scaffolds led to in vivo VSMC lineage specification and matrix deposition inside the scaffolds. In summary, we established integration-free patient-specific hiPSCs from peripheral blood mononuclear cells, derived CVPCs through directed lineage specification, and developed an advanced scaffold for these progenitor cells to further differentiate in vivo into VSMCs and regenerate vascular tissue in a subcutaneous implantation model. This study has established an efficient patient-specific approach towards in vivo regeneration of vascular tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cell-free collagen-based scaffolds used for making blood vessels in cardiovascular surgery.

PubMed

Akhmedov, Sh D; Afanas'ev, S A; Egorova, M V; Andreev, S L; Ivanov, A V; Rogovskaia, Yu V; Usov, V Yu; Shvedov, A N; Steinhoff, G

2012-01-01

The present article deals with the technology of obtaining decellularized cell-free collagen-based scaffolds from arterial vessels and surgical assessment of the possibility of experimentally implanting them into the blood system of laboratory animals for experimental purposes. The study was performed on arterial vessels (n=60) and fragments of the human internal thoracic artery (n=20). Described herein is a method of obtaining a connective-tissue matrix of a blood vessel by means of vessel's perfusion for 2-3 hours with detergent solutions. Cell-free collagen-based conduits were implanted to a total of ten dogs. After the operation, the blood flow remained functional. The anastomoses established turned out to be leak-proof and the acellular vessels were able to withstand the haemodynamic load of the arterial blood flow.

Molecular parallels between neural and vascular development.

PubMed

Eichmann, Anne; Thomas, Jean-Léon

2013-01-01

The human central nervous system (CNS) features a network of ~400 miles of blood vessels that receives >20% of the body's cardiac output and uses most of its blood glucose. Many human diseases, including stroke, retinopathy, and cancer, are associated with the biology of CNS blood vessels. These vessels originate from extrinsic cell populations, including endothelial cells and pericytes that colonize the CNS and interact with glia and neurons to establish the blood-brain barrier and control cerebrovascular exchanges. Neurovascular interactions also play important roles in adult neurogenic niches, which harbor a unique population of neural stem cells that are intimately associated with blood vessels. We here review the cellular and molecular mechanisms required to establish the CNS vascular network, with a special focus on neurovascular interactions and the functions of vascular endothelial growth factors.

Transplantation of autologous peripheral blood mononuclear cells in the subarachnoid space for amyotrophic lateral sclerosis: a safety analysis of 14 patients

PubMed Central

Li, Xiao-yan; Liang, Zhan-hua; Han, Chao; Wei, Wen-juan; Song, Chun-li; Zhou, Li-na; Liu, Yang; Li, Ying; Ji, Xiao-fei; Liu, Jing

2017-01-01

There is a small amount of clinical data regarding the safety and feasibility of autologous peripheral blood mononuclear cell transplantation into the subarachnoid space for the treatment of amyotrophic lateral sclerosis. The objectives of this retrospective study were to assess the safety and efficacy of peripheral blood mononuclear cell transplantation in 14 amyotrophic lateral sclerosis patients to provide more objective data for future clinical trials. After stem cell mobilization and collection, autologous peripheral blood mononuclear cells (1 × 109) were isolated and directly transplanted into the subarachnoid space of amyotrophic lateral sclerosis patients. The primary outcome measure was incidence of adverse events. Secondary outcome measures were electromyography 1 week before operation and 4 weeks after operation, Functional Independence Measurement, Berg Balance Scale, and Dysarthria Assessment Scale 1 week preoperatively and 1, 2, 4 and 12 weeks postoperatively. There was no immediate or delayed transplant-related cytotoxicity. The number of leukocytes, serum alanine aminotransferase and creatinine levels, and body temperature were within the normal ranges. Radiographic evaluation showed no serious transplant-related adverse events. Muscle strength grade, results of Functional Independence Measurement, Berg Balance Scale, and Dysarthria Assessment Scale were not significantly different before and after treatment. These findings suggest that peripheral blood mononuclear cell transplantation into the subarachnoid space for the treatment of amyotrophic lateral sclerosis is safe, but its therapeutic effect is not remarkable. Thus, a large-sample investigation is needed to assess its efficacy further. PMID:28469667

Platelets and their interactions with other immune cells

PubMed Central

Lam, Fong W.; Vijayan, K. Vinod; Rumbaut, Rolando E.

2015-01-01

Platelets are anucleate blood cells, long known to be critically involved in hemostasis and thrombosis. In addition to their role in blood clots, increasing evidence reveals significant roles for platelets in inflammation and immunity. However, the notion that platelets represent immune cells is not broadly recognized in the field of Physiology. This manuscript reviews the role of platelets in inflammation and immune responses, and highlights their interactions with other immune cells, including examples of major functional consequences of these interactions. PMID:26140718

Solid lipid nanoparticles as a vehicle for brain-targeted drug delivery: two new strategies of functionalization with apolipoprotein E

NASA Astrophysics Data System (ADS)

Rute Neves, Ana; Fontes Queiroz, Joana; Weksler, Babette; Romero, Ignacio A.; Couraud, Pierre-Olivier; Reis, Salette

2015-12-01

Nanotechnology can be an important tool to improve the permeability of some drugs for the blood-brain barrier. In this work we created a new system to enter the brain by functionalizing solid lipid nanoparticles with apolipoprotein E, aiming to enhance their binding to low-density lipoprotein receptors on the blood-brain barrier endothelial cells. Solid lipid nanoparticles were successfully functionalized with apolipoprotein E using two distinct strategies that took advantage of the strong interaction between biotin and avidin. Transmission electron microscopy images revealed spherical nanoparticles, and dynamic light scattering gave a Z-average under 200 nm, a polydispersity index below 0.2, and a zeta potential between -10 mV and -15 mV. The functionalization of solid lipid nanoparticles with apolipoprotein E was demonstrated by infrared spectroscopy and fluorimetric assays. In vitro cytotoxic effects were evaluated by MTT and LDH assays in the human cerebral microvascular endothelial cells (hCMEC/D3) cell line, a human blood-brain barrier model, and revealed no toxicity up to 1.5 mg ml-1 over 4 h of incubation. The brain permeability was evaluated in transwell devices with hCMEC/D3 monolayers, and a 1.5-fold increment in barrier transit was verified for functionalized nanoparticles when compared with non-functionalized ones. The results suggested that these novel apolipoprotein E-functionalized nanoparticles resulted in dynamic stable systems capable of being used for an improved and specialized brain delivery of drugs through the blood-brain barrier.

Comparative Study on Compositions and Functional Properties of Porcine, Chicken and Duck Blood

PubMed Central

2017-01-01

Hematological, chemical and functional characteristics of porcine, chicken and duck blood were evaluated. A porcine blood sample showed the most abundant red blood cell, hemoglobin concentration, packed cell volume and plasma protein content as well as its freeze-dried blood possessed the highest contents of protein, fat, Cu and Cr with the highest percentage of heme iron (p<0.05). Unlike porcine blood, chicken blood showed a well balance in some essential amino acids, specifically for a higher isoleucine content (p<0.05). Furthermore, it possessed the highest contents of carbohydrate, Zn and non-heme iron (p<0.05). The most rapid response to form a strong gel, especially at 70°C and 80°C, was found in chicken blood, followed by duck and porcine blood, respectively. The result of emulsion activity index (EAI) and emulsion stability index (ESI) at the low protein concentration indicated that chicken blood had the most superior emulsion properties (p<0.05). Regarding duck blood, it exhibited the highest content of Mg and Mn (p<0.05). Moreover, duck blood had similar foaming properties to porcine blood in which they showed higher values than chicken blood (p<0.05). Specific characteristics of blood were therefore diminished by animal species in which this information could be used as food supplementation or product development based on their potential applications. PMID:28515647

Paul Ehrlich and the Early History of Granulocytes.

PubMed

Kay, A Barry

2016-08-01

Paul Ehrlich's techniques, published between 1879 and 1880, for staining blood films using coal tar dyes, and his method of differential blood cell counting, ended years of speculation regarding the classification of white cells. Acidic and basic dyes had allowed him to recognize eosinophil and basophil granules, respectively, work that was a direct continuation of his discovery of the tissue mast cell described in his doctoral thesis. Ehrlich went on to develop neutral dyes that identified epsilon granules in neutrophils ("cells with polymorphous nuclei"). He also speculated, for the most part correctly, on the formation, function, and fate of blood neutrophils and eosinophils. Before Ehrlich, a number of important observations had been made on white cells and their role in health and disease. Among the most notable were William Hewson's studies of blood and lymph; the early descriptions of leukemia by Alfred Donné, John Hughes Bennett, Rudolf Virchow, and others; as well as seminal observations on inflammation by William Addison, Friedrich von Recklinghausen, and Julius Cohnheim. Eosinophils were almost certainly recognized previously by others. In 1846, Thomas Wharton Jones (1808-1891) described "granule blood-cells" in several species including humans. The term "granule cell" had also been used by Julius Vogel (1814-1880), who had previously observed similar cells in inflammatory exudates. Vogel, in turn, was aware of the work of Gottlieb Gluge (1812-1898), who had observed "compound inflammatory globules" in pus and serum that resembled eosinophils. Almost 20 years before Ehrlich developed his staining methods, Max Johann Schultze (1825-1874) performed functional experiments on fine and coarse granular cells using a warm stage microscopic technique and showed they had amoeboid movement and phagocytic abilities. Despite these earlier observations, it was Ehrlich's use of stains that heralded the modern era of studies of leukocyte biology and pathology.

Folic acid functionalized surface highlights 5-methylcytosine-genomic content within circulating tumor cells.

PubMed

Malara, Natalia; Coluccio, Maria Laura; Limongi, Tania; Asande, Monica; Trunzo, Valentina; Cojoc, Gheorghe; Raso, Cinzia; Candeloro, Patrizio; Perozziello, Gerardo; Raimondo, Raffaella; De Vitis, Stefania; Roveda, Laura; Renne, Maria; Prati, Ubaldo; Mollace, Vincenzo; Di Fabrizio, Enzo

2014-11-12

Although the detection of methylated cell free DNA represents one of the most promising approaches for relapse risk assessment in cancer patients, the low concentration of cell-free circulating DNA constitutes the biggest obstacle in the development of DNA methylation-based biomarkers from blood. This paper describes a method for the measurement of genomic methylation content directly on circulating tumor cells (CTC), which could be used to deceive the aforementioned problem. Since CTC are disease related blood-based biomarkers, they result essential to monitor tumor's stadiation, therapy, and early relapsing lesions. Within surface's bio-functionalization and cell's isolation procedure standardization, the presented approach reveals a singular ability to detect high 5-methylcytosine CTC-subset content in the whole CTC compound, by choosing folic acid (FA) as transducer molecule. Sensitivity and specificity, calculated for FA functionalized surface (FA-surface), result respectively on about 83% and 60%. FA-surface, allowing the detection and characterization of early metastatic dissemination, provides a unique advance in the comprehension of tumors progression and dissemination confirming the presence of CTC and its association with high risk of relapse. This functionalized surface identifying and quantifying high 5-methylcytosine CTC-subset content into the patient's blood lead significant progress in cancer risk assessment, also providing a novel therapeutic strategy. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reversibility of peripheral blood leukocyte phenotypic and functional changes after exposure to and withdrawal from tofacitinib, a Janus kinase inhibitor, in healthy volunteers.

PubMed

Weinhold, Kent J; Bukowski, Jack F; Brennan, Todd V; Noveck, Robert J; Staats, Janet S; Lin, Liwen; Stempora, Linda; Hammond, Constance; Wouters, Ann; Mojcik, Christopher F; Cheng, John; Collinge, Mark; Jesson, Michael I; Hazra, Anasuya; Biswas, Pinaki; Lan, Shuping; Clark, James D; Hodge, Jennifer A

2018-06-01

This study evaluated the short-term effects of tofacitinib treatment on peripheral blood leukocyte phenotype and function, and the reversibility of any such effects following treatment withdrawal in healthy volunteers. Cytomegalovirus (CMV)-seropositive subjects received oral tofacitinib 10 mg twice daily for 4 weeks and were followed for 4 weeks after drug withdrawal. There were slight increases in total lymphocyte and total T-cell counts during tofacitinib treatment, and B-cell counts increased by up to 26%. There were no significant changes in granulocyte or monocyte counts, or granulocyte function. Naïve and central memory T-cell counts increased during treatment, while all subsets of activated T cells were decreased by up to 69%. T-cell subsets other than effector memory cluster of differentiation (CD)4+, activated naïve CD4+ and effector CD8+ T-cell counts and B-cell counts, normalized 4 weeks after withdrawal. Following ex vivo activation, measures of CMV-specific T-cell responses, and antigen non-specific T-cell-mediated cytotoxicity and interferon (IFN)-γ production, decreased slightly. These T-cell functional changes were most pronounced at Day 15, partially normalized while still on tofacitinib and returned to baseline after drug withdrawal. Total natural killer (NK)-cell counts decreased by 33%, returning towards baseline after drug withdrawal. NK-cell function decreased during tofacitinib treatment, but without a consistent time course across measured parameters. However, markers of NK-cell-mediated cytotoxicity, antibody-dependent cellular cytotoxicity and IFN-γ production were decreased up to 42% 1 month after drug withdrawal. CMV DNA was not detectable in whole blood, and there were no cases of herpes zoster reactivation. No new safety concerns arose. In conclusion, the effect of short-term tofacitinib treatment on leukocyte composition and function in healthy CMV+ volunteers is modest and largely reversible 4 weeks after withdrawal. Copyright © 2018 Pfizer Inc. Published by Elsevier Inc. All rights reserved.

A smart core-sheath nanofiber that captures and releases red blood cells from the blood.

PubMed

Shi, Q; Hou, J; Zhao, C; Xin, Z; Jin, J; Li, C; Wong, S-C; Yin, J

2016-01-28

A smart core-sheath nanofiber for non-adherent cell capture and release is demonstrated. The nanofibers are fabricated by single-spinneret electrospinning of poly(N-isopropylacrylamide) (PNIPAAm), polycaprolactone (PCL) and nattokinase (NK) solution blends. The self-assembly of PNIPAAm and PCL blends during the electrospinning generates the core-sheath PCL/PNIPAAm nanofibers with PNIPAAm as the sheath. The PNIPAAm-based core-sheath nanofibers are switchable between hydrophobicity and hydrophilicity with temperature change and enhance stability in the blood. When the nanofibers come in contact with blood, the NK is released from the nanofibers to resist platelet adhesion on the nanofiber surface, facilitating the direct capture and isolation of red blood cells (RBCs) from the blood above phase-transition temperature of PNIPAAm. Meanwhile, the captured RBCs are readily released from the nanofibers with temperature stimuli in an undamaged manner. The release efficiency of up to 100% is obtained while maintaining cellular integrity and function. This work presents promising nanofibers to effectively capture non-adherent cells and release for subsequent molecular analysis and diagnosis of single cells.

MicroRNAs enriched in hematopoietic stem cells differentially regulate long-term hematopoietic output.

PubMed

O'Connell, Ryan M; Chaudhuri, Aadel A; Rao, Dinesh S; Gibson, William S J; Balazs, Alejandro B; Baltimore, David

2010-08-10

The production of blood cells depends on a rare hematopoietic stem-cell (HSC) population, but the molecular mechanisms underlying HSC biology remain incompletely understood. Here, we identify a subset of microRNAs (miRNAs) that is enriched in HSCs compared with other bone-marrow cells. An in vivo gain-of-function screen found that three of these miRNAs conferred a competitive advantage to engrafting hematopoietic cells, whereas other HSC miRNAs attenuated production of blood cells. Overexpression of the most advantageous miRNA, miR-125b, caused a dose-dependent myeloproliferative disorder that progressed to a lethal myeloid leukemia in mice and also enhanced hematopoietic engraftment in human immune system mice. Our study identifies an evolutionarily conserved subset of miRNAs that is expressed in HSCs and functions to modulate hematopoietic output.

Efficient and safe gene delivery to human corneal endothelium using magnetic nanoparticles.

PubMed

Czugala, Marta; Mykhaylyk, Olga; Böhler, Philip; Onderka, Jasmine; Stork, Björn; Wesselborg, Sebastian; Kruse, Friedrich E; Plank, Christian; Singer, Bernhard B; Fuchsluger, Thomas A

2016-07-01

To develop a safe and efficient method for targeted, anti-apoptotic gene therapy of corneal endothelial cells (CECs). Magnetofection (MF), a combination of lipofection with magnetic nanoparticles (MNPs; PEI-Mag2, SO-Mag5, PalD1-Mag1), was tested in human CECs and in explanted human corneas. Effects on cell viability and function were investigated. Immunocompatibility was assessed in human peripheral blood mononuclear cells. Silica iron-oxide MNPs (SO-Mag5) combined with X-tremeGENE-HP achieved high transfection efficiency in human CECs and explanted human corneas, without altering cell viability or function. Magnetofection caused no immunomodulatory effects in human peripheral blood mononuclear cells. Magnetofection with anti-apoptotic P35 gene effectively blocked apoptosis in CECs. Magnetofection is a promising tool for gene therapy of corneal endothelial cells with potential for targeted on-site delivery.

Whole Blood Activation Results in Enhanced Detection of T Cell and Monocyte Cytokine Production by Flow Cytometry

NASA Technical Reports Server (NTRS)

Sams, Clarence F.; Crucian, Brian E.

2001-01-01

An excellent monitor of the immune balance of peripheral circulating cells is to determine their cytokine production patterns in response to stimuli. Using flow cytometry a positive identification of cytokine producing cells in a mixed culture may be achieved. Recently, the ability to assess cytokine production following a wholeblood activation culture has been described. We compared whole blood culture to standard PBMC culture and determined the individual cytokine secretion patterns for both T cells and monocytes via flow cytometry. For T cells cytokine assessment following PMA +ionomycin activation: (1) a significantly greater percentages of T cells producing IFNgamma and IL-2 were observed following whole-blood culture; (2) altered T cell cytokine production kinetics were observed by varying whole blood culture times. In addition, a four-color cytometric analysis was used to allow accurate phenotyping and quantitation of cytokine producing lymphocyte populations. Using this technique we found IFNgamma production to be significantly elevated in the CD3+/CD8+ T cell population as compared to the CD3+/CD8- population following five hours of whole blood activation. Conversely, IL-2 and IL-10 production were significantly elevated in the CD3+/CD8- T cell population as compared to the CD3+/CD8+ population. Monocyte cytokine production was assessed in both culture systems following LPS activation for 24 hours. A three-color flow cytometric was used to assess two cytokines in conjunction with CD 14. The cytokine pairs used for analysis were IL-1a/IL-12, and IL-10ITNFa. Nearly all monocytes were stimulated to produce IL-1a, IL-12 and TNFalpha equally well in both culture systems. Monocyte production of IL-10 was significantly elevated following whole blood culture as compared to PBMC culture. IL-12 producing monocytes appeared to be a distinct subpopulation of the IL-1a producing set, whereas IL-10 and TNFa producing monocytes were largely mutually exclusive. IL-10 and TNFa producing monocytes may represent functionally different monocyte subsets with distinct functions. Whole blood culture eliminates the need to purify cell populations prior to culture and may have significant utility for the routine monitoring of the cytokine balances of the peripheral blood T cell and monocyte populations. In addition, there are distinct advantages to performing whole-blood (WB) activation as compared to PBMC activation. These advantages would include retaining all various cell-cell interactions as well as any soluble factors present in serum that influence cell activation. It is likely that the altered cytokine production observed following whole blood culture more accurately represents the in-vivo immune balance.

Peripheral blood film review. The demise of the eyecount leukocyte differential.

PubMed

Pierre, Robert V

2002-03-01

The automated hematology analyzer with CBC and differential results has replaced the traditional manual or individual assay methods for hematologic parameters and the eyecount leukocyte differential as the initial screening and detection system for hematologic abnormalities in modern hospitals and clinics. The traditional review of all automated hematology instrument results by preparation, staining, and microscopic examination of a blood film has disappeared in most institutions. The reasons are the more accurate detection of specimens with distributional or morphologic abnormalities by the instruments than by the traditional eyecount method. The opportunity for a clinician to request a microscopic examination of a blood film, whether or not it is flagged, must be preserved, because the clinician's knowledge of the patient's history, physical findings, and current or prior therapy may indicate review to discover an abnormality that may not have been apparent from the instrument results alone. There has also been a dramatic reduction of the numbers of medical technologists and technicians in medical laboratories. Automation of the CBC and differential counts has reduced the number of technologists needed for performance of these tests. But other factors have had a negative effect, such as the necessity to reduce costs. Consolidation of hematology and chemistry laboratories in core laboratories may produce savings in labor costs, but may also create problems of creating and maintaining areas of expertise, such as hematologic morphology, because of the cross-training required and the necessity of personnel to do all things. This article suggests and documents a number of measures that can be infinity stituted by the laboratory and by clinicians to reduce the number of eyecount differentials and blood film reviews that need to be performed. The first effort is to convince clinicians that valid data exist that confirm that a policy of allowing the laboratory to initiate blood film review based on findings of the CBC and automated differential is a more sensitive and accurate method of detecting patients with blood film abnormalities than routine blood film review of all specimens by technologists. Clinicians need to recognize that daily differential results or differentials at intervals of less than a week are not medically necessary in most patients. The laboratory, however, must provide opportunities for the clinician to request differentials at any time for specific medical reasons. The laboratory must establish the validity of screening criteria for detection of distribution and morphologic abnormalities of leukocytes by clinical correlation studies or adopt criteria established by laboratories with the same instrumentation and which have conducted clinical evaluations. A final observation on the eyecount differential is that it was the only way to identify cell types and their relative proportion for nearly 100 years. Cells were identified by their shape, intracellular structures, and staining characteristics. Many studies were able eventually to correlate some aspect of each cell type's function with their morphologic appearance. It has also been learned that the bone marrow is the source of production of most circulating cells and a great deal of the controls of cell production and release into the peripheral blood have been learned. But leukocytes have many functions, almost none of which are performed in the peripheral blood. The peripheral blood is mainly a conduit from the bone marrow to the tissues where the leukocytes perform their function in the case of the neutrophils and monocytes. It is mainly a recirculation and redistribution system for lymphocytes that usually receive their instructions from antigen processing cells in the tissues and allow these modified cells to home to sites where their functions occur. Cellular morphology and staining characteristics tell little about the maturation stage and functional capabilities of leukocytes. One cannot tell the difference between a band and a segmented neutrophil or whether a lymphocyte is a T or B cell on the conventional eyecount differential. One cannot tell the mature granulocyte of a patient with chronic myeloid leukemia from a normal mature neutrophil. Increasingly, techniques are being developed to identify better the maturation stages of cells and association with specific functional capabilities by flow cytometric techniques. The neoplastic nature of some normal-appearing leukocytes can be identified by techniques, such as fluorescent in situ hybridization. With the rapid advances in many approachs to understand the nature and functional capability of leukocytes, the eyecount differential with the traditional Romanowsky stain may be past the apogee of its ascent and beginning its trip into history along with the hemocytometer counting chamber and the Sahli pipet. The development and implementation of new laboratory cornerstone techniques for diagnosis of hematologic disease are eagerly awaited. On the other hand, the red cells and platelets exist to function in the peripheral blood. More emphasis is needed in the development of automated methods of determining the nature and functional capabilities of these true blood cells as part of the CBC.

High red blood cell nitric oxide synthase activation is not associated with improved vascular function and red blood cell deformability in sickle cell anaemia.

PubMed

Grau, Marijke; Mozar, Anaïs; Charlot, Keyne; Lamarre, Yann; Weyel, Linda; Suhr, Frank; Collins, Bianca; Jumet, Stéphane; Hardy-Dessources, Marie-Dominique; Romana, Marc; Lemonne, Nathalie; Etienne-Julan, Maryse; Antoine-Jonville, Sophie; Bloch, Wilhelm; Connes, Philippe

2015-03-01

Human red blood cells (RBC) express an active and functional endothelial-like nitric oxide (NO) synthase (RBC-NOS). We report studies on RBC-NOS activity in sickle cell anaemia (SCA), a genetic disease characterized by decreased RBC deformability and vascular dysfunction. Total RBC-NOS content was not significantly different in SCA patients compared to healthy controls; however, using phosphorylated RBC-NOS-Ser(1177) as a marker, RBC-NOS activation was higher in SCA patients as a consequence of the greater activation of Akt (phosphorylated Akt-Ser(473) ). The higher RBC-NOS activation in SCA led to higher levels of S-nitrosylated α- and β-spectrins, and greater RBC nitrite and nitrotyrosine levels compared to healthy controls. Plasma nitrite content was not different between the two groups. Laser Doppler flowmetric experiments demonstrated blunted microcirculatory NO-dependent response under hyperthermia in SCA patients. RBC deformability, measured by ektacytometry, was reduced in SCA in contrast to healthy individuals, and pre-shearing RBC in vitro did not improve deformability despite an increase of RBC-NOS activation. RBC-NOS activation is high in freshly drawn blood from SCA patients, resulting in high amounts of NO produced by RBC. However, this does not result in improved RBC deformability and vascular function: higher RBC-NO is not sufficient to counterbalance the enhanced oxidative stress in SCA. © 2014 John Wiley & Sons Ltd.

[Stimulation of cell cultures recovery after cryopreservation by the cattle cord blood FRACTION (below 5 kDa) or Actovegin].

PubMed

Gulevskiĭ, A K; Trifonova, A V; Lavrik, A A

2013-01-01

The capacities of the cattle cord blood low-molecular fraction (below 5 kDa) and Actovegin (the vealer blood fraction (below 5 kDa)) for recovering functions of cell cultures after cryopreservation compared. Their influence proliferation of the flozen-thawed cell cultures, certain stages of their growth, cell attachment, rate of cell spreading, and mitotic regiment has been studied. Both the cord blood low-molecular fraction and Actovegin were shown to stimulate growth of the cell cultures after cryopreservation more efficiently at the concentration of 224 μg/ml. However, despite the stimulating effect discovered, their application did not bring proliferative indices on the 1st passage after cryopreservation to the values of the native culture. The effects of the cord blood low-molecular fraction and Actovegin on the human fibroblast culture were identical by the following parameters: cell attachment, rates of cell spreading and proliferation. In culture BHK-21 clone 13/04 the efficiency of Actovegin was low, while the cord blood low-molecular fraction has a conspicuous stimulating effect on its adhesion and proliferation. The investigations carried out can serve as a basis for the development of regenerative media containing the cattle cord blood low-molecular fraction (below 5 kDa) or Actovegin as active components at the concentration of 224 μg/ml with the purpose of fast recovery of culture prolifetative properties after cryopreservation.

An overview of the role of microparticles/microvesicles in blood components: Are they clinically beneficial or harmful?

PubMed

Burnouf, Thierry; Chou, Ming-Li; Goubran, Hadi; Cognasse, Fabrice; Garraud, Olivier; Seghatchian, Jerard

2015-10-01

Blood cells and tissues generate heterogeneous populations of cell-derived vesicles, ranging from approximately 50 nm to 1 µm in diameter. Under normal physiological conditions and as an essential part of an energy-dependent natural process, microparticles (MPs) are continuously shed into the circulation from membranes of all viable cells such as megakaryocytes, platelets, red blood cells, white blood cells and endothelial cells. MP shedding can also be triggered by pathological activation of inflammatory processes and activation of coagulation or complement systems, or even by shear stress in the circulation. Structurally, MPs have a bilayered phospholipid structure exposing coagulant-active phosphatidylserine and expressing various membrane receptors, and they serve as cell-to-cell shuttles for bioactive molecules such as lipids, growth factors, microRNAs, and mitochondria. It was established that ex vivo processing of blood into its components, involving centrifugation, processing by various apheresis procedures, leucoreduction, pathogen reduction, and finally storage in different media and different types of blood bags, can impact MP generation and content. This is mostly due to exposure of the collected blood to anticoagulant/storage media and due to shear stresses or activation, contact with artificial surfaces, or exposure to various leucocyte-removal filters and pathogen-reduction treatments. Such artificially generated MPs, which are added to the original pool of MPs collected from the donor, may exhibit specific functional characteristics, as MPs are not an inert element of blood components. Not surprisingly, MPs' roles and functionality are therefore increasingly seen to be fully relevant to the field of transfusion medicine, and as a parameter of blood safety that must be considered in haemovigilance programmes. Continual advancements in assessment methods of MPs and storage lesions are gradually leading to a better understanding of the impacts of blood collection on MP generation, while clinical research should clarify links of MPs with transfusion reactions and certain clinical disorders. Harmonization and consensus in sampling protocols, sample handling and processing, and assessment methods are needed to achieve consensual interpretations. This review focuses on the role of MPs as an essential laboratory tool and as a most effective player in transfusion science and medicine and in health and disease. Copyright © 2015 Elsevier Ltd. All rights reserved.

Pathogen inactivation treatment of plasma and platelet concentrates and their predicted functionality in massive transfusion protocols.

PubMed

Arbaeen, Ahmad F; Schubert, Peter; Serrano, Katherine; Carter, Cedric J; Culibrk, Brankica; Devine, Dana V

2017-05-01

Trauma transfusion packages for hemorrhage control consist of red blood cells, plasma, and platelets at a set ratio. Although pathogen reduction improves the transfusion safety of platelet and plasma units, there is an associated reduction in quality. This study aimed to investigate the impact of riboflavin/ultraviolet light-treated plasma or platelets in transfusion trauma packages composed of red blood cell, plasma, and platelet units in a ratio of 1:1:1 in vitro by modeling transfusion scenarios for trauma patients and assessing function by rotational thromboelastometry. Pathogen-reduced or untreated plasma and buffy coat platelet concentrate units produced in plasma were used in different combinations with red blood cells in trauma transfusion packages. After reconstitution of these packages with hemodiluted blood, the hemostatic functionality was analyzed by rotational thromboelastometry. Hemostatic profiles of pathogen-inactivated buffy coat platelet concentrate and plasma indicated decreased activity compared with their respective controls. Reconstitution of hemodiluted blood (hematocrit = 20%) with packages that contained treated or nontreated components resulted in increased alpha and maximum clot firmness and enhanced clot-formation time. Simulating transfusion scenarios based on 30% blood replacement with a transfusion trauma package resulted in a nonsignificant difference in rotational thromboelastometry parameters between packages containing treated and nontreated blood components (p ≥ 0.05). Effects of pathogen inactivation treatment were evident when the trauma package percentage was 50% or greater and contained both pathogen inactivation-treated plasma and buffy coat platelet concentrate. Rotational thromboelastometry investigations suggest that there is relatively little impact of pathogen inactivation treatment on whole blood clot formation unless large amounts of treated components are used. © 2017 AABB.

Platelets generated from human embryonic stem cells are functional in vitro and in the microcirculation of living mice

PubMed Central

Lu, Shi-Jiang; Li, Feng; Yin, Hong; Feng, Qiang; Kimbrel, Erin A; Hahm, Eunsil; Thon, Jonathan N; Wang, Wei; Italiano, Joseph E; Cho, Jaehyung; Lanza, Robert

2011-01-01

Platelets play an essential role in hemostasis and atherothrombosis. Owing to their short storage time, there is constant demand for this life-saving blood component. In this study, we report that it is feasible to generate functional megakaryocytes and platelets from human embryonic stem cells (hESCs) on a large scale. Differential-interference contrast and electron microscopy analyses showed that ultrastructural and morphological features of hESC-derived platelets were indistinguishable from those of normal blood platelets. In functional assays, hESC-derived platelets responded to thrombin stimulation, formed microaggregates, and facilitated clot formation/retraction in vitro. Live cell microscopy demonstrated that hESC-platelets formed lamellipodia and filopodia in response to thrombin activation, and tethered to each other as observed in normal blood. Using real-time intravital imaging with high-speed video microscopy, we have also shown that hESC-derived platelets contribute to developing thrombi at sites of laser-induced vascular injury in mice, providing the first evidence for in vivo functionality of hESC-derived platelets. These results represent an important step toward generating an unlimited supply of platelets for transfusion. Since platelets contain no genetic material, they are ideal candidates for early clinical translation involving human pluripotent stem cells. PMID:21221130

Engineering of functional, perfusable 3D microvascular networks on a chip.

PubMed

Kim, Sudong; Lee, Hyunjae; Chung, Minhwan; Jeon, Noo Li

2013-04-21

Generating perfusable 3D microvessels in vitro is an important goal for tissue engineering, as well as for reliable modelling of blood vessel function. To date, in vitro blood vessel models have not been able to accurately reproduce the dynamics and responses of endothelial cells to grow perfusable and functional 3D vascular networks. Here we describe a microfluidic-based platform whereby we model natural cellular programs found during normal development and angiogenesis to form perfusable networks of intact 3D microvessels as well as tumor vasculatures based on the spatially controlled co-culture of endothelial cells with stromal fibroblasts, pericytes or cancer cells. The microvessels possess the characteristic morphological and biochemical markers of in vivo blood vessels, and exhibit strong barrier function and long-term stability. An open, unobstructed microvasculature allows the delivery of nutrients, chemical compounds, biomolecules and cell suspensions, as well as flow-induced mechanical stimuli into the luminal space of the endothelium, and exhibits faithful responses to physiological shear stress as demonstrated by cytoskeleton rearrangement and increased nitric oxide synthesis. This simple and versatile platform provides a wide range of applications in vascular physiology studies as well as in developing vascularized organ-on-a-chip and human disease models for pharmaceutical screening.

Hematopoiesis and hematopoietic organs in arthropods.

PubMed

Grigorian, Melina; Hartenstein, Volker

2013-03-01

Hemocytes (blood cells) are motile cells that move throughout the extracellular space and that exist in all clades of the animal kingdom. Hemocytes play an important role in shaping the extracellular environment and in the immune response. Developmentally, hemocytes are closely related to the epithelial cells lining the vascular system (endothelia) and the body cavity (mesothelia). In vertebrates and insects, common progenitors, called hemangioblasts, give rise to the endothelia and blood cells. In the adult animal, many differentiated hemocytes seem to retain the ability to proliferate; however, in most cases investigated closely, the bulk of hemocyte proliferation takes place in specialized hematopoietic organs. Hematopoietic organs provide an environment where undifferentiated blood stem cells are able to self-renew, and at the same time generate offspring that differentiate into different blood cell types. Hematopoiesis in vertebrates, taking place in the bone marrow, has been subject to intensive research by immunologists and stem cell biologists. Much less is known about blood cell formation in invertebrate animals. In this review, we will survey structural and functional properties of invertebrate hematopoietic organs, with a main focus on insects and other arthropod taxa. We will then discuss similarities, at the molecular and structural level, that are apparent when comparing the development of blood cells in hematopoietic organs of vertebrates and arthropods. Our comparative review is intended to elucidate aspects of the biology of blood stem cells that are more easily missed when focusing on one or a few model species.

A Combined Negative and Positive Enrichment Assay for Cancer Cells Isolation and Purification.

PubMed

Cheng, Boran; Wang, Shuyi; Chen, Yuanyuan; Fang, Yuan; Chen, Fangfang; Wang, Zhenmeng; Xiong, Bin

2016-02-01

Cancer cells that detach from solid tumor and circulate in the peripheral blood (CTCs) have been considered as a new "biomarker" for the detection and characterization of cancers. However, isolating and detecting cancer cells from the cancer patient peripheral blood have been technically challenging, owing to the small sub-population of CTCs (a few to hundreds per milliliter). Here we demonstrate a simple and efficient cancer cells isolation and purification method. A biocompatible and surface roughness controllable TiO2 nanofilm was deposited onto a glass slide to achieve enhanced topographic interactions with nanoscale cellular surface components, again, anti-CD45 (a leukocyte common antigen) and anti-EpCAM (epithelial cell adhesion molecule) were then coated onto the surface of the nanofilm for advance depletion of white blood cells (WBCs) and specific isolation of CTCs, respectively. Comparing to the conventional positive enrichment technology, this method exhibited excellent biocompatibility and equally high capture efficiency. Moreover, the maximum number of background cells (WBCs) was removed, and viable and functional cancer cells were isolated with high purity. Utilizing the horizontally packed TiO2 nanofilm improved pure CTC-capture through combining cell-capture-agent and cancer cell-preferred nanoscale topography, which represented a new method capable of obtaining biologically functional CTCs for subsequent molecular analysis. © The Author(s) 2014.

Differential white cell count by centrifugal microfluidics.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sommer, Gregory Jon; Tentori, Augusto M.; Schaff, Ulrich Y.

We present a method for counting white blood cells that is uniquely compatible with centrifugation based microfluidics. Blood is deposited on top of one or more layers of density media within a microfluidic disk. Spinning the disk causes the cell populations within whole blood to settle through the media, reaching an equilibrium based on the density of each cell type. Separation and fluorescence measurement of cell types stained with a DNA dye is demonstrated using this technique. The integrated signal from bands of fluorescent microspheres is shown to be proportional to their initial concentration in suspension. Among the current generationmore » of medical diagnostics are devices based on the principle of centrifuging a CD sized disk functionalized with microfluidics. These portable 'lab on a disk' devices are capable of conducting multiple assays directly from a blood sample, embodied by platforms developed by Gyros, Samsung, and Abaxis. [1,2] However, no centrifugal platform to date includes a differential white blood cell count, which is an important metric complimentary to diagnostic assays. Measuring the differential white blood cell count (the relative fraction of granulocytes, lymphocytes, and monocytes) is a standard medical diagnostic technique useful for identifying sepsis, leukemia, AIDS, radiation exposure, and a host of other conditions that affect the immune system. Several methods exist for measuring the relative white blood cell count including flow cytometry, electrical impedance, and visual identification from a stained drop of blood under a microscope. However, none of these methods is easily incorporated into a centrifugal microfluidic diagnostic platform.« less

IL‐12 and IL‐15 induce the expression of CXCR6 and CD49a on peripheral natural killer cells

PubMed Central

Hydes, Theresa; Noll, Angela; Salinas‐Riester, Gabriela; Abuhilal, Mohammed; Armstrong, Thomas; Hamady, Zaed; Primrose, John; Takhar, Arjun; Walter, Lutz

2017-01-01

Abstract Introduction Murine hepatic NK cells exhibit adaptive features, with liver‐specific adhesion molecules CXCR6 and CD49a acting as surface markers. Methods We investigated human liver‐resident CXCR6+ and CD49a+ NK cells using RNA sequencing, flow cytometry, and functional analysis. We further assessed the role of cytokines in generating NK cells with these phenotypes from the peripheral blood. Results Hepatic CD49a+ NK cells could be induced using cytokines and produce high quantities of IFNγ and TNFα, in contrast to hepatic CXCR6+ NK cells. RNA sequencing of liver‐resident CXCR6+ NK cells confirmed a tolerant immature phenotype with reduced expression of markers associated with maturity and cytotoxicity. Liver‐resident double‐positive CXCR6 + CD49a+ hepatic NK cells are immature but maintain high expression of Th1 cytokines as observed for single‐positive CD49a+ NK cells. We show that stimulation with activating cytokines can readily induce upregulation of both CD49a and CXCR6 on NK cells in the peripheral blood. In particular, IL‐12 and IL‐15 can generate CXCR6 + CD49a+ NK cells in vitro from NK cells isolated from the peripheral blood, with comparable phenotypic and functional features to liver‐resident CD49a+ NK cells, including enhanced IFNγ and NKG2C expression. Conclusion IL‐12 and IL‐15 may be key for generating NK cells with a tissue‐homing phenotype and strong Th1 cytokine profile in the blood, and links peripheral activation of NK cells with tissue‐homing. These findings may have important therapeutic implications for immunotherapy of chronic liver disease. PMID:28952190

IL-12 and IL-15 induce the expression of CXCR6 and CD49a on peripheral natural killer cells.

PubMed

Hydes, Theresa; Noll, Angela; Salinas-Riester, Gabriela; Abuhilal, Mohammed; Armstrong, Thomas; Hamady, Zaed; Primrose, John; Takhar, Arjun; Walter, Lutz; Khakoo, Salim I

2018-03-01

Murine hepatic NK cells exhibit adaptive features, with liver-specific adhesion molecules CXCR6 and CD49a acting as surface markers. We investigated human liver-resident CXCR6+ and CD49a+ NK cells using RNA sequencing, flow cytometry, and functional analysis. We further assessed the role of cytokines in generating NK cells with these phenotypes from the peripheral blood. Hepatic CD49a+ NK cells could be induced using cytokines and produce high quantities of IFNγ and TNFα, in contrast to hepatic CXCR6+ NK cells. RNA sequencing of liver-resident CXCR6+ NK cells confirmed a tolerant immature phenotype with reduced expression of markers associated with maturity and cytotoxicity. Liver-resident double-positive CXCR6 + CD49a+ hepatic NK cells are immature but maintain high expression of Th1 cytokines as observed for single-positive CD49a+ NK cells. We show that stimulation with activating cytokines can readily induce upregulation of both CD49a and CXCR6 on NK cells in the peripheral blood. In particular, IL-12 and IL-15 can generate CXCR6 + CD49a+ NK cells in vitro from NK cells isolated from the peripheral blood, with comparable phenotypic and functional features to liver-resident CD49a+ NK cells, including enhanced IFNγ and NKG2C expression. IL-12 and IL-15 may be key for generating NK cells with a tissue-homing phenotype and strong Th1 cytokine profile in the blood, and links peripheral activation of NK cells with tissue-homing. These findings may have important therapeutic implications for immunotherapy of chronic liver disease. © 2017 The Authors. Immunity, Inflammation and Disease Published by John Wiley & Sons Ltd.

Blood flow structure in patients with coronary heart disease

NASA Astrophysics Data System (ADS)

Malinova, Lidia I.; Simonenko, Georgy V.; Denisova, Tatyana P.; Tuchin, Valery V.

2007-05-01

Blood flow structure was studied by PC integrated video camera with following slide by slide analysis. Volumetric blood flow velocity was supporting on constant level (1 ml/h). Silicone tube of diameter comparable with coronary arteries diameter was used as vessel model. Cell-cell interactions were studied under glucose and anticoagulants influence. Increased adhesiveness of blood cells to tube walls was revealed in patient with coronary heart disease (CHD) compare to practically healthy persons (PHP). In patients with stable angina pectoris of high functional class and patients with AMI shear stress resistant erythrocyte aggregates were predominating in blood flow structure up to microclots formation. Clotting and erythrocytes aggregation increase as response to glucose solution injection, sharply defined in patients with CHD. Heparin injection (10 000 ED) increased linear blood flow velocity both in patients with CHD and PHP. After compare our results with other author's data we can consider that method used in our study is sensible enough to investigate blood flow structure violations in patients with CHD and PHP. Several differences of cell-cell interaction in flow under glucose and anticoagulant influence were found out in patients with CHD and PHP.

The controversial origin of pericytes during angiogenesis - Implications for cell-based therapeutic angiogenesis and cell-based therapies.

PubMed

Blocki, Anna; Beyer, Sebastian; Jung, Friedrich; Raghunath, Michael

2018-01-01

Pericytes reside within the basement membrane of small vessels and are often in direct cellular contact with endothelial cells, fulfilling important functions during blood vessel formation and homeostasis. Recently, these pericytes have been also identified as mesenchymal stem cells. Mesenchymal stem cells, and especially their specialized subpopulation of pericytes, represent promising candidates for therapeutic angiogenesis applications, and have already been widely applied in pre-clinical and clinical trials. However, cell-based therapies of ischemic diseases (especially of myocardial infarction) have not resulted in significant long-term improvement. Interestingly, pericytes from a hematopoietic origin were observed in embryonic skin and a pericyte sub-population expressing leukocyte and monocyte markers was described during adult angiogenesis in vivo. Since mesenchymal stem cells do not express hematopoietic markers, the latter cell type might represent an alternative pericyte population relevant to angiogenesis. Therefore, we sourced blood-derived angiogenic cells (BDACs) from monocytes that closely resembled hematopoietic pericytes, which had only been observed in vivo thus far. BDACs displayed many pericytic features and exhibited enhanced revascularization and functional tissue regeneration in a pre-clinical model of critical limb ischemia. Comparison between BDACs and mesenchymal pericytes indicated that BDACs (while resembling hematopoietic pericytes) enhanced early stages of angiogenesis, such as endothelial cell sprouting. In contrast, mesenchymal pericytes were responsible for blood vessel maturation and homeostasis, while reducing endothelial sprouting.Since the formation of new blood vessels is crucial during therapeutic angiogenesis or during integration of implants into the host tissue, hematopoietic pericytes (and therefore BDACs) might offer an advantageous addition or even an alternative for cell-based therapies.

Liquid biopsy in cancer patients: advances in capturing viable CTCs for functional studies using the EPISPOT assay.

PubMed

Alix-Panabières, Catherine; Pantel, Klaus

2015-01-01

Circulating tumor cells (CTCs) in the blood of cancer patients have received increasing attention as new diagnostic tool enabling 'liquid biopsies'. In contrast to the wealth of descriptive studies demonstrating the clinical relevance of CTCs as biomarkers, the extremely low concentration of CTCs in the peripheral blood of most cancer patients challenges further functional studies. This article discusses the current possibilities to enrich and, in particular, detect viable CTCs with emphasis on the EPithelial ImmunoSPOT technology. This functional assay detects viable CTCs at the single-cell level and has been used on hundreds of patients with different tumor types including epithelial tumors (breast, prostate and colon cancer) and melanomas. Moreover, the article summarizes recent advances in the in vitro and in vivo expansion of CTCs from cancer patients. These functional analyses will contribute to identifying the biological properties of metastatic cells and reveal new therapeutic targets against disseminating cancer cells.

Human endothelial progenitor cells-derived exosomes accelerate cutaneous wound healing in diabetic rats by promoting endothelial function.

PubMed

Li, Xiaocong; Jiang, Chunyu; Zhao, Jungong

2016-08-01

Wound healing is deeply dependent on neovascularization to restore blood flow. The neovascularization of endothelial progenitor cells (EPCs) through paracrine secretion has been reported in various tissue repair models. Exosomes, key components of cell paracrine mechanism, have been rarely reported in wound healing. Exosomes were isolated from the media of EPCs obtained from human umbilical cord blood. Diabetic rats wound model was established and treated with exosomes. The in vitro effects of exosomes on the proliferation, migration and angiogenic tubule formation of endothelial cells were investigated. We revealed that human umbilical cord blood EPCs derived exosomes transplantation could accelerate cutaneous wound healing in diabetic rats. We also showed that exosomes enhanced the proliferation, migration and tube formation of vascular endothelial cells in vitro. Furthermore, we found that endothelial cells stimulated with these exosomes would increase expression of angiogenesis-related molecules, including FGF-1, VEGFA, VEGFR-2, ANG-1, E-selectin, CXCL-16, eNOS and IL-8. Taken together, our findings indicated that EPCs-derived exosomes facilitate wound healing by positively modulating vascular endothelial cells function. Copyright © 2016 Elsevier Inc. All rights reserved.

[Proliferation and osteogenic differentiation of mesenchymal stem cells in hydrogels of human blood plasma].

PubMed

Linero, Itali M; Doncel, Adriana; Chaparro, Orlando

2014-01-01

The use of mesenchymal stem cells in clinical practice has increased considerably in the last decade because they play a supporting role in the processes of tissue repair and regeneration, becoming the main tool of cell therapy for the treatment of diseases functionally affecting bone and cartilage tissue . To evaluate in vitro the proliferative and osteogenic differentiation ability of mesenchymal stem cells derived from human adipose tissue in a blood plasma hydrogel. Mesenchymal stem cells were obtained from human adipose tissue explants and characterized by flow cytometry. Their multipotentiality was demonstrated by their ability to differentiate to adipogenic and osteogenic lineages. Cell proliferation and osteogenic differentiation ability of the cells cultured in blood plasma hydrogels were also evaluated. Mesenchymal stem cells derived from human adipose tissue growing in human blood plasma hydrogels showed a pattern of proliferation similar to that of the cells cultured in monolayer and also maintained their ability to differentiate to osteogenic lineage. Human blood plasma hydrogels are a suitable support for proliferation and osteogenic differentiation of mesenchymal stem cells derived from human adipose tissue and provides a substrate that is autologous, biocompatible, reabsorbable, easy to use, potentially injectable and economic, which could be used as a successful strategy for the management and clinical application of cell therapy in regenerative medicine.

Application of a clot-based assay to measure the procoagulant activity of stored allogeneic red blood cell concentrates

PubMed Central

Wannez, Adeline; Bailly, Nicolas; Alpan, Lutfiye; Gheldof, Damien; Douxfils, Jonathan; Deneys, Véronique; Bihin, Benoît; Chatelain, Bernard; Dogné, Jean-Michel; Chatelain, Christian; Mullier, François

2018-01-01

Background Thrombotic effects are possible complications of red blood cell transfusion. The generation and accumulation of procoagulant red blood cell extracellular vesicles during storage may play an important role in these thrombotic effects. The objective of this study was to assess the value of a simple phospholipid-dependent clot-based assay (STA®-Procoag-PPL) to estimate the procoagulant activity of stored red blood cells and changes in this activity during storage of the blood component. Materials and methods Extracellular vesicles from 12 red blood cell concentrates were isolated at 13 storage time-points and characterised by quantitative and functional methods: the degree of haemolysis (direct spectrophotometry), the quantification and determination of cellular origin (flow cytometry) and the procoagulant activity (thrombin generation and STA®-Procoag-PPL assays) were assessed. Results The mean clotting time of extracellular vesicles isolated from red blood cell concentrates decreased from 117.2±3.6 sec on the day of collection to 33.8±1.3 sec at the end of the storage period. This illustrates the phospholipid-dependent procoagulant activity of these extracellular vesicles, as confirmed by thrombin generation. Results of the peak of thrombin and the STA®-Procoag-PPL were well correlated (partial r=−0.41. p<0.001). In parallel, an exponential increase of the number of red blood cell-derived extracellular vesicles from 1,779/μL to 218,451/μL was observed. Discussion The STA®-Procoag-PPL is a potentially useful technique for assessing the procoagulant activity of a red blood cell concentrate. PMID:28287378

Drosophila hematopoiesis under normal conditions and in response to immune stress.

PubMed

Letourneau, Manon; Lapraz, Francois; Sharma, Anurag; Vanzo, Nathalie; Waltzer, Lucas; Crozatier, Michèle

2016-11-01

The emergence of hematopoietic progenitors and their differentiation into various highly specialized blood cell types constitute a finely tuned process. Unveiling the genetic cascades that control blood cell progenitor fate and understanding how they are modulated in response to environmental changes are two major challenges in the field of hematopoiesis. In the last 20 years, many studies have established important functional analogies between blood cell development in vertebrates and in the fruit fly, Drosophila melanogaster. Thereby, Drosophila has emerged as a powerful genetic model for studying mechanisms that control hematopoiesis during normal development or in pathological situations. Moreover, recent advances in Drosophila have highlighted how intricate cell communication networks and microenvironmental cues regulate blood cell homeostasis. They have also revealed the striking plasticity of Drosophila mature blood cells and the presence of different sites of hematopoiesis in the larva. This review provides an overview of Drosophila hematopoiesis during development and summarizes our current knowledge on the molecular processes controlling larval hematopoiesis, both under normal conditions and in response to an immune challenge, such as wasp parasitism. © 2016 Federation of European Biochemical Societies.

GPU-accelerated Red Blood Cells Simulations with Transport Dissipative Particle Dynamics.

PubMed

Blumers, Ansel L; Tang, Yu-Hang; Li, Zhen; Li, Xuejin; Karniadakis, George E

2017-08-01

Mesoscopic numerical simulations provide a unique approach for the quantification of the chemical influences on red blood cell functionalities. The transport Dissipative Particles Dynamics (tDPD) method can lead to such effective multiscale simulations due to its ability to simultaneously capture mesoscopic advection, diffusion, and reaction. In this paper, we present a GPU-accelerated red blood cell simulation package based on a tDPD adaptation of our red blood cell model, which can correctly recover the cell membrane viscosity, elasticity, bending stiffness, and cross-membrane chemical transport. The package essentially processes all computational workloads in parallel by GPU, and it incorporates multi-stream scheduling and non-blocking MPI communications to improve inter-node scalability. Our code is validated for accuracy and compared against the CPU counterpart for speed. Strong scaling and weak scaling are also presented to characterizes scalability. We observe a speedup of 10.1 on one GPU over all 16 cores within a single node, and a weak scaling efficiency of 91% across 256 nodes. The program enables quick-turnaround and high-throughput numerical simulations for investigating chemical-driven red blood cell phenomena and disorders.

Red blood cells promote survival and cell cycle progression of human peripheral blood T cells independently of CD58/LFA-3 and heme compounds.

PubMed

Fonseca, Ana Mafalda; Pereira, Carlos Filipe; Porto, Graça; Arosa, Fernando A

2003-07-01

Red blood cells (RBC) are known to modulate T cell proliferation and function possibly through downregulation of oxidative stress. By examining parameters of activation, division, and cell death in vitro, we show evidence that the increase in survival afforded by RBC is due to the maintenance of the proliferative capacity of the activated T cells. We also show that the CD3+CD8+ T cell subset was preferentially expanded and rescued from apoptosis both in bulk peripheral blood lymphocyte cultures and with highly purified CD8+ T cells. The ability of RBC to induce survival of dividing T cells was not affected by blocking the CD58/CD2 interaction. Moreover, addition of hemoglobin, heme or protoporphyrin IX to cultures of activated T cells did not reproduce the effect of intact RBC. Considering that RBC circulate throughout the body, they could play a biological role in the modulation of T cell differentiation and survival in places of active cell division. Neither CD58 nor the heme compounds studied seem to play a direct relevant role in the modulation of T cell survival.

Bone Marrow Transplantation Results in Human Donor Blood Cells Acquiring and Displaying Mouse Recipient Class I MHC and CD45 Antigens on Their Surface

PubMed Central

Yamanaka, Nobuko; Wong, Christine J.; Gertsenstein, Marina; Casper, Robert F.; Nagy, Andras; Rogers, Ian M.

2009-01-01

Background Mouse models of human disease are invaluable for determining the differentiation ability and functional capacity of stem cells. The best example is bone marrow transplants for studies of hematopoietic stem cells. For organ studies, the interpretation of the data can be difficult as transdifferentiation, cell fusion or surface antigen transfer (trogocytosis) can be misinterpreted as differentiation. These events have not been investigated in hematopoietic stem cell transplant models. Methodology/Principal Findings In this study we investigated fusion and trogocytosis involving blood cells during bone marrow transplantation using a xenograft model. We report that using a standard SCID repopulating assay almost 100% of the human donor cells appear as hybrid blood cells containing both mouse and human surface antigens. Conclusion/Significance Hybrid cells are not the result of cell-cell fusion events but appear to be due to efficient surface antigen transfer, a process referred to as trogocytosis. Antigen transfer appears to be non-random and includes all donor cells regardless of sub-type. We also demonstrate that irradiation preconditioning enhances the frequency of hybrid cells and that trogocytosis is evident in non-blood cells in chimera mice. PMID:20046883

Optimal occlusion uniformly partitions red blood cells fluxes within a microvascular network

PubMed Central

Tu, Shenyinying; Liu, Yu-Hsiu; Savage, Van M.; Hsiai, Tzung K.; Roper, Marcus

2017-01-01

In animals, gas exchange between blood and tissues occurs in narrow vessels, whose diameter is comparable to that of a red blood cell. Red blood cells must deform to squeeze through these narrow vessels, transiently blocking or occluding the vessels they pass through. Although the dynamics of vessel occlusion have been studied extensively, it remains an open question why microvessels need to be so narrow. We study occlusive dynamics within a model microvascular network: the embryonic zebrafish trunk. We show that pressure feedbacks created when red blood cells enter the finest vessels of the trunk act together to uniformly partition red blood cells through the microvasculature. Using mathematical models as well as direct observation, we show that these occlusive feedbacks are tuned throughout the trunk network to prevent the vessels closest to the heart from short-circuiting the network. Thus occlusion is linked with another open question of microvascular function: how are red blood cells delivered at the same rate to each micro-vessel? Our analysis shows that tuning of occlusive feedbacks increase the total dissipation within the network by a factor of 11, showing that uniformity of flows rather than minimization of transport costs may be prioritized by the microvascular network. PMID:29244812

Therapeutic PD-L1 and LAG-3 blockade rapidly clears established blood-stage Plasmodium infection

PubMed Central

Butler, Noah S.; Moebius, Jacqueline; Pewe, Lecia L.; Traore, Boubacar; Doumbo, Ogobara K.; Tygrett, Lorraine T.; Waldschmidt, Thomas J.; Crompton, Peter D.; Harty, John T.

2011-01-01

Plasmodium infection of erythrocytes induces clinical malaria. Parasite-specific CD4+ T cells correlate with reduced parasite burdens and severity of human malaria, and are required to control blood-stage infection in mice. However, the characteristics of CD4+ T cells that determine protection or parasite persistence remain unknown. Here we show that P. falciparum infection of humans increased expression of an inhibitory receptor (PD-1) associated with T cell dysfunction. In vivo blockade of PD-L1 and LAG-3 restored CD4+ T cell function, amplified T follicular helper cell and germinal center B cell and plasmablast numbers, enhanced protective antibodies and rapidly cleared blood-stage malaria in mice. Thus, chronic malaria drives specific T cell dysfunction, which can be rescued to enhance parasite control using inhibitory therapies. PMID:22157630

Nanotube antibody biosensor arrays for the detection of circulating breast cancer cells

NASA Astrophysics Data System (ADS)

Shao, Ning; Wickstrom, Eric; Panchapakesan, Balaji

2008-11-01

Recent reports have shown that nanoscale electronic devices can be used to detect a change in electrical properties when receptor proteins bind to their corresponding antibodies functionalized on the surface of the device, in extracts from as few as ten lysed tumor cells. We hypothesized that nanotube-antibody devices could sensitively and specifically detect entire live cancer cells. We report for the first time a single nanotube field effect transistor array, functionalized with IGF1R-specific and Her2-specific antibodies, which exhibits highly sensitive and selective sensing of live, intact MCF7 and BT474 human breast cancer cells in human blood. Those two cell lines both overexpress IGF1R and Her2, at different levels. Single or small bundle of nanotube devices that were functionalized with IGF1R-specific or Her2-specific antibodies showed 60% decreases in conductivity upon interaction with BT474 or MCF7 breast cancer cells in two µl drops of blood. Control experiments with non-specific antibodies or with MCF10A control breast cells produced a less than 5% decrease in electrical conductivity, illustrating the high sensitivity for whole cell binding by these single nanotube-antibody devices. We postulate that the free energy change due to multiple simultaneous cell-antibody binding events exerted stress along the nanotube surface, decreasing its electrical conductivity due to an increase in band gap. Because the free energy change upon cell-antibody binding, the stress exerted on the nanotube, and the change in conductivity are specific to a specific antigen-antibody interaction; these properties might be used as a fingerprint for the molecular sensing of circulating cancer cells. From optical microscopy observations during sensing, it appears that the binding of a single cell to a single nanotube field effect transistor produced the change in electrical conductivity. Thus we report a nanoscale oncometer with single cell sensitivity with a diameter 1000 times smaller than a cancer cell that functions in a drop of fresh blood.

Brain and Retinal Pericytes: Origin, Function and Role

PubMed Central

Trost, Andrea; Lange, Simona; Schroedl, Falk; Bruckner, Daniela; Motloch, Karolina A.; Bogner, Barbara; Kaser-Eichberger, Alexandra; Strohmaier, Clemens; Runge, Christian; Aigner, Ludwig; Rivera, Francisco J.; Reitsamer, Herbert A.

2016-01-01

Pericytes are specialized mural cells located at the abluminal surface of capillary blood vessels, embedded within the basement membrane. In the vascular network these multifunctional cells fulfil diverse functions, which are indispensable for proper homoeostasis. They serve as microvascular stabilizers, are potential regulators of microvascular blood flow and have a central role in angiogenesis, as they for example regulate endothelial cell proliferation. Furthermore, pericytes, as part of the neurovascular unit, are a major component of the blood-retina/brain barrier. CNS pericytes are a heterogenic cell population derived from mesodermal and neuro-ectodermal germ layers acting as modulators of stromal and niche environmental properties. In addition, they display multipotent differentiation potential making them an intriguing target for regenerative therapies. Pericyte-deficiencies can be cause or consequence of many kinds of diseases. In diabetes, for instance, pericyte-loss is a severe pathological process in diabetic retinopathy (DR) with detrimental consequences for eye sight in millions of patients. In this review, we provide an overview of our current understanding of CNS pericyte origin and function, with a special focus on the retina in the healthy and diseased. Finally, we highlight the role of pericytes in de- and regenerative processes. PMID:26869887

Automatic tracking of red blood cells in micro channels using OpenCV

NASA Astrophysics Data System (ADS)

Rodrigues, Vânia; Rodrigues, Pedro J.; Pereira, Ana I.; Lima, Rui

2013-10-01

The present study aims to developan automatic method able to track red blood cells (RBCs) trajectories flowing through a microchannel using the Open Source Computer Vision (OpenCV). The developed method is based on optical flux calculation assisted by the maximization of the template-matching product. The experimental results show a good functional performance of this method.

A dynamic in vivo-like organotypic blood-brain barrier model to probe metastatic brain tumors

NASA Astrophysics Data System (ADS)

Xu, Hui; Li, Zhongyu; Yu, Yue; Sizdahkhani, Saman; Ho, Winson S.; Yin, Fangchao; Wang, Li; Zhu, Guoli; Zhang, Min; Jiang, Lei; Zhuang, Zhengping; Qin, Jianhua

2016-11-01

The blood-brain barrier (BBB) restricts the uptake of many neuro-therapeutic molecules, presenting a formidable hurdle to drug development in brain diseases. We proposed a new and dynamic in vivo-like three-dimensional microfluidic system that replicates the key structural, functional and mechanical properties of the blood-brain barrier in vivo. Multiple factors in this system work synergistically to accentuate BBB-specific attributes-permitting the analysis of complex organ-level responses in both normal and pathological microenvironments in brain tumors. The complex BBB microenvironment is reproduced in this system via physical cell-cell interaction, vascular mechanical cues and cell migration. This model possesses the unique capability to examine brain metastasis of human lung, breast and melanoma cells and their therapeutic responses to chemotherapy. The results suggest that the interactions between cancer cells and astrocytes in BBB microenvironment might affect the ability of malignant brain tumors to traverse between brain and vascular compartments. Furthermore, quantification of spatially resolved barrier functions exists within a single assay, providing a versatile and valuable platform for pharmaceutical development, drug testing and neuroscientific research.

Modulation of early functional recovery of Achilles tendon to bone unit after transection by BPC 157 and methylprednisolone.

PubMed

Krivic, A; Majerovic, M; Jelic, I; Seiwerth, S; Sikiric, P

2008-05-01

In the presented study we compared the effect of stable peptide BPC 157 and methylprednisolone on early functional recovery after Achilles tendon to bone transection in a rat model before collagen healing started. Surgical transection of the right Achilles tendon to bone area was performed in seventy two Wistar Albino male rats. Healing Achilles tendon edges were harvested at days 1-4 following the transection. Using Achilles functional index (AFI), myeloperoxidase activity, histological inflammatory cell influx and vascular index early functional recovery was evaluated. Agents (stable peptide BPC 157 10 microg methylprednisolone 5 mg, normal saline 5 ml) were given alone (/kg b.w., intraperitoneally, once daily, first 30 min after surgery, last 24 h before analysis). Control group received normal saline 5 ml/kg. BPC 157 improved functional recovery (AFI values increased at all time points, p <0.05) by anti-inflammatory (decreased myeloperoxidase (MPO) activity and histological inflammatory cell influx, p <0.05) and increased new blood vessel formation (increased vascular index, p <0.05). Methyprednisolone decreased MPO activity and histological inflammatory cell influx, (p <0.05) but also decreased new blood vessel formation and did not affect early functional recovery. Stable peptide BPC 157 with combined anti-inflammatory action and induction of early new blood vessel formation facilitates early functional recovery in Achilles tendon to bone healing.

Profiling MHC II immunopeptidome of blood-stage malaria reveals that cDC1 control the functionality of parasite-specific CD4 T cells.

PubMed

Draheim, Marion; Wlodarczyk, Myriam F; Crozat, Karine; Saliou, Jean-Michel; Alayi, Tchilabalo Dilezitoko; Tomavo, Stanislas; Hassan, Ali; Salvioni, Anna; Demarta-Gatsi, Claudia; Sidney, John; Sette, Alessandro; Dalod, Marc; Berry, Antoine; Silvie, Olivier; Blanchard, Nicolas

2017-11-01

In malaria, CD4 Th1 and T follicular helper (T FH ) cells are important for controlling parasite growth, but Th1 cells also contribute to immunopathology. Moreover, various regulatory CD4 T-cell subsets are critical to hamper pathology. Yet the antigen-presenting cells controlling Th functionality, as well as the antigens recognized by CD4 T cells, are largely unknown. Here, we characterize the MHC II immunopeptidome presented by DC during blood-stage malaria in mice. We establish the immunodominance hierarchy of 14 MHC II ligands derived from conserved parasite proteins. Immunodominance is shaped differently whether blood stage is preceded or not by liver stage, but the same ETRAMP-specific dominant response develops in both contexts. In naïve mice and at the onset of cerebral malaria, CD8α + dendritic cells (cDC1) are superior to other DC subsets for MHC II presentation of the ETRAMP epitope. Using in vivo depletion of cDC1, we show that cDC1 promote parasite-specific Th1 cells and inhibit the development of IL-10 + CD4 T cells. This work profiles the P. berghei blood-stage MHC II immunopeptidome, highlights the potency of cDC1 to present malaria antigens on MHC II, and reveals a major role for cDC1 in regulating malaria-specific CD4 T-cell responses. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

Maternal body-mass index and cord blood circulating endothelial colony-forming cells.

PubMed

Moreno-Luna, Rafael; Muñoz-Hernandez, Rocio; Lin, Ruei-Zeng; Miranda, Maria L; Vallejo-Vaz, Antonio J; Stiefel, Pablo; Praena-Fernández, Juan M; Bernal-Bermejo, Jose; Jimenez-Jimenez, Luis M; Villar, Jose; Melero-Martin, Juan M

2014-03-01

Endothelial colony-forming cells (ECFCs) are a subset of circulating endothelial progenitor cells that are particularly abundant in umbilical cord blood. We sought to determine whether ECFC abundance in cord blood is associated with maternal body-mass index (BMI) in nonpathologic pregnancies. We measured the level of ECFCs in the cord blood of neonates (n = 27) born from non-obese healthy mothers with nonpathologic pregnancies and examined whether ECFC abundance correlated with maternal BMI. We also examined the effect of maternal BMI on ECFC phenotype and function using angiogenic and vasculogenic assays. We observed variation in ECFC abundance among subjects and found a positive correlation between prepregnancy maternal BMI and ECFC content (r = 0.51, P = .007), which was independent of other obstetric factors. Despite this variation, ECFC phenotype and functionality were deemed normal and highly similar between subjects with maternal BMI <25 kg/m(2) and BMI between 25-30 kg/m(2), including the ability to form vascular networks in vivo. This study underlines the need to consider maternal BMI as a potential confounding factor for cord blood levels of ECFCs in future comparative studies between healthy and pathologic pregnancies. Copyright © 2014 Mosby, Inc. All rights reserved.

Interdialytic ambulatory blood pressure in patients with intradialytic hypertension.

PubMed

Van Buren, Peter N; Toto, Robert; Inrig, Jula K

2012-01-01

Hypertension is common in hemodialysis patients and contributes to this population's high risk for cardiovascular morbidity and mortality. Patients with intradialytic hypertension, or increases in blood pressure during hemodialysis, have been shown to have the highest risk for these outcomes. The purpose of this review is to describe new findings that shed light on the epidemiology and pathophysiology of intradialytic hypertension and discuss how a better understanding of these mechanisms may lead to improved blood pressure management and outcomes in hemodialysis patients. Our laboratory demonstrated that intradialytic hypertension occurs at least sporadically in most hemodialysis patients, but in 25% of patients it occurs in over 31% of their hemodialysis treatments. We also identified that, compared with hemodialysis patients without intradialytic hypertension, those with intradialytic hypertension have worse endothelial cell function and have higher interdialytic ambulatory blood pressure. Pilot study data show that carvedilol reduces the frequency of intradialytic hypertension and improves endothelial cell dysfunction. Intradialytic hypertension is associated with increased morbidity and mortality, impaired endothelial cell function, and higher overall blood pressure burden. Further investigation is required to determine whether interventions aimed at preventing or treating intradialytic hypertension improve long-term outcomes.

Dissecting gene expression at the blood-brain barrier

PubMed Central

Huntley, Melanie A.; Bien-Ly, Nga; Daneman, Richard; Watts, Ryan J.

2014-01-01

The availability of genome-wide expression data for the blood-brain barrier is an invaluable resource that has recently enabled the discovery of several genes and pathways involved in the development and maintenance of the blood-brain barrier, particularly in rodent models. The broad distribution of published data sets represents a viable starting point for the molecular dissection of the blood-brain barrier and will further direct the discovery of novel mechanisms of blood-brain barrier formation and function. Technical advances in purifying brain endothelial cells, the key cell that forms the critical barrier, have allowed for greater specificity in gene expression comparisons with other central nervous system cell types, and more systematic characterizations of the molecular composition of the blood-brain barrier. Nevertheless, our understanding of how the blood-brain barrier changes during aging and disease is underrepresented. Blood-brain barrier data sets from a wider range of experimental paradigms and species, including invertebrates and primates, would be invaluable for investigating the function and evolution of the blood-brain barrier. Newer technologies in gene expression profiling, such as RNA-sequencing, now allow for finer resolution of transcriptomic changes, including isoform specificity and RNA-editing. As our field continues to utilize more advanced expression profiling in its ongoing efforts to elucidate the blood-brain barrier, including in disease and drug delivery, we will continue to see rapid advances in our understanding of the molecular mediators of barrier biology. We predict that the recently published data sets, combined with forthcoming genomic and proteomic blood-brain barrier data sets, will continue to fuel the molecular genetic revolution of blood-brain barrier biology. PMID:25414634

An integrated microfluidic chip system for single-cell secretion profiling of rare circulating tumor cells.

PubMed

Deng, Yuliang; Zhang, Yu; Sun, Shuai; Wang, Zhihua; Wang, Minjiao; Yu, Beiqin; Czajkowsky, Daniel M; Liu, Bingya; Li, Yan; Wei, Wei; Shi, Qihui

2014-12-16

Genetic and transcriptional profiling, as well as surface marker identification of single circulating tumor cells (CTCs) have been demonstrated. However, quantitatively profiling of functional proteins at single CTC resolution has not yet been achieved, owing to the limited purity of the isolated CTC populations and a lack of single-cell proteomic approaches to handle and analyze rare CTCs. Here, we develop an integrated microfluidic system specifically designed for streamlining isolation, purification and single-cell secretomic profiling of CTCs from whole blood. Key to this platform is the use of photocleavable ssDNA-encoded antibody conjugates to enable a highly purified CTC population with <75 'contaminated' blood cells. An enhanced poly-L-lysine barcode pattern is created on the single-cell barcode chip for efficient capture rare CTC cells in microchambers for subsequent secreted protein profiling. This system was extensively evaluated and optimized with EpCAM-positive HCT116 cells seeded into whole blood. Patient blood samples were employed to assess the utility of the system for isolation, purification and single-cell secretion profiling of CTCs. The CTCs present in patient blood samples exhibit highly heterogeneous secretion profile of IL-8 and VEGF. The numbers of secreting CTCs are found not in accordance with CTC enumeration based on immunostaining in the parallel experiments.

An Integrated Microfluidic Chip System for Single-Cell Secretion Profiling of Rare Circulating Tumor Cells

PubMed Central

Deng, Yuliang; Zhang, Yu; Sun, Shuai; Wang, Zhihua; Wang, Minjiao; Yu, Beiqin; Czajkowsky, Daniel M.; Liu, Bingya; Li, Yan; Wei, Wei; Shi, Qihui

2014-01-01

Genetic and transcriptional profiling, as well as surface marker identification of single circulating tumor cells (CTCs) have been demonstrated. However, quantitatively profiling of functional proteins at single CTC resolution has not yet been achieved, owing to the limited purity of the isolated CTC populations and a lack of single-cell proteomic approaches to handle and analyze rare CTCs. Here, we develop an integrated microfluidic system specifically designed for streamlining isolation, purification and single-cell secretomic profiling of CTCs from whole blood. Key to this platform is the use of photocleavable ssDNA-encoded antibody conjugates to enable a highly purified CTC population with <75 ‘contaminated' blood cells. An enhanced poly-L-lysine barcode pattern is created on the single-cell barcode chip for efficient capture rare CTC cells in microchambers for subsequent secreted protein profiling. This system was extensively evaluated and optimized with EpCAM-positive HCT116 cells seeded into whole blood. Patient blood samples were employed to assess the utility of the system for isolation, purification and single-cell secretion profiling of CTCs. The CTCs present in patient blood samples exhibit highly heterogeneous secretion profile of IL-8 and VEGF. The numbers of secreting CTCs are found not in accordance with CTC enumeration based on immunostaining in the parallel experiments. PMID:25511131

CFD analysis of multiphase blood flow within aorta and its thoracic branches of patient with coarctation of aorta using multiphase Euler - Euler approach

NASA Astrophysics Data System (ADS)

Ostrowski, Z.; Melka, B.; Adamczyk, W.; Rojczyk, M.; Golda, A.; Nowak, A. J.

2016-09-01

In the research a numerical Computational Fluid Dynamics (CFD) model of the pulsatile blood flow was created and analyzed. A real geometry of aorta and its thoracic branches of 8-year old patient diagnosed with a congenital heart defect - coarctation of aorta was used. The inlet boundary condition were implemented as the User Define Function according to measured values of volumetric blood flow. The blood flow was treated as multiphase: plasma, set as the primary fluid phase, was dominant with volume fraction of 0.585 and morphological elements of blood were treated in Euler-Euler approach as dispersed phases (with 90% Red Blood Cells and White Blood Cells as remaining solid volume fraction).

Coxsackie- and adenovirus receptor (CAR) is expressed in lymphatic vessels in human skin and affects lymphatic endothelial cell function in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vigl, Benjamin; Zgraggen, Claudia; Rehman, Nadia

Lymphatic vessels play an important role in tissue fluid homeostasis, intestinal fat absorption and immunosurveillance. Furthermore, they are involved in pathologic conditions, such as tumor cell metastasis and chronic inflammation. In comparison to blood vessels, the molecular phenotype of lymphatic vessels is less well characterized. Performing comparative gene expression analysis we have recently found that coxsackie- and adenovirus receptor (CAR) is significantly more highly expressed in cultured human, skin-derived lymphatic endothelial cells (LECs), as compared to blood vascular endothelial cells. Here, we have confirmed these results at the protein level, using Western blot and FACS analysis. Immunofluorescence performed on humanmore » skin confirmed that CAR is expressed at detectable levels in lymphatic vessels, but not in blood vessels. To address the functional significance of CAR expression, we modulated CAR expression levels in cultured LECs in vitro by siRNA- and vector-based transfection approaches. Functional assays performed with the transfected cells revealed that CAR is involved in distinct cellular processes in LECs, such as cell adhesion, migration, tube formation and the control of vascular permeability. In contrast, no effect of CAR on LEC proliferation was observed. Overall, our data suggest that CAR stabilizes LEC-LEC interactions in the skin and may contribute to lymphatic vessel integrity.« less

Novel silicon microchannels device for use in red blood cell deformability studies

NASA Astrophysics Data System (ADS)

Zheng, Xiao-Lin; Liao, Yan-Jian; Zhang, Wen-Xian

2001-10-01

Currently, a number of techniques are used to access cell deformability. We study a novel silicon microchannels device for use in red blood cell deformability. The channels are produced in silicon substrate using microengineering technology. The microgrooves formed in the surface of a single-crystal silicon substrate. They were converted to channels by tightly covering them with an optical flat glass plate. An array of flow channels (number 950 in parallel) have typical dimensions of 5 micrometers width X 5.5 Xm depth, and 30 micrometers length. There the RBC's are forced to pass through channels. Thus, the microchannels are used to simulate human blood capillaries. It provides a specific measurement of individual cell in terms of both flow velocity profile and an index of cell volume while the cell flow through the channels. It dominates the complex cellular flow behavior, such as, the viscosity of whole blood is a nonlinear function of shear rate, index of filtration, etc.

Longitudinal Assessment of Vascular Function With Sunitinib in Patients With Metastatic Renal Cell Carcinoma.

PubMed

Catino, Anna B; Hubbard, Rebecca A; Chirinos, Julio A; Townsend, Ray; Keefe, Stephen; Haas, Naomi B; Puzanov, Igor; Fang, James C; Agarwal, Neeraj; Hyman, David; Smith, Amanda M; Gordon, Mary; Plappert, Theodore; Englefield, Virginia; Narayan, Vivek; Ewer, Steven; ElAmm, Chantal; Lenihan, Daniel; Ky, Bonnie

2018-03-01

Sunitinib, used widely in metastatic renal cell carcinoma, can result in hypertension, left ventricular dysfunction, and heart failure. However, the relationships between vascular function and cardiac dysfunction with sunitinib are poorly understood. In a multicenter prospective study of 84 metastatic renal cell carcinoma patients, echocardiography, arterial tonometry, and BNP (B-type natriuretic peptide) measures were performed at baseline and at 3.5, 15, and 33 weeks after sunitinib initiation, correlating with sunitinib cycles 1, 3, and 6. Mean change in vascular function parameters and 95% confidence intervals were calculated. Linear regression models were used to estimate associations between vascular function and left ventricular ejection fraction, longitudinal strain, diastolic function (E/e'), and BNP. After 3.5 weeks of sunitinib, mean systolic blood pressure increased by 9.5 mm Hg (95% confidence interval, 2.0-17.1; P =0.02) and diastolic blood pressure by 7.2 mm Hg (95% confidence interval, 4.3-10.0; P <0.001) across all participants. Sunitinib resulted in increases in large artery stiffness (carotid-femoral pulse wave velocity) and resistive load (total peripheral resistance and arterial elastance; all P <0.05) and changes in pulsatile load (total arterial compliance and wave reflection). There were no statistically significant associations between vascular function and systolic dysfunction (left ventricular ejection fraction and longitudinal strain). However, baseline total peripheral resistance, arterial elastance, and aortic impedance were associated with worsening diastolic function and filling pressures over time. In patients with metastatic renal cell carcinoma, sunitinib resulted in early, significant increases in blood pressure, arterial stiffness, and resistive and pulsatile load within 3.5 weeks of treatment. Baseline vascular function parameters were associated with worsening diastolic but not systolic function. © 2018 American Heart Association, Inc.

Elevated red blood cell distribution width is associated with liver function tests in patients with primary hepatocellular carcinoma.

PubMed

Wei, Ting-Ting; Tang, Qing-Qin; Qin, Bao-Dong; Ma, Ning; Wang, Li-Li; Zhou, Lin; Zhong, Ren-Qian

2016-11-25

Red blood cell distribution width (RDW), a routinely tested parameter of the complete blood count (CBC), has been reported to be increased in various cancers and correlated with the patients' clinical characteristics. However, the significance of RDW in primary hepatocellular carcinoma (pHCC) is largely unknown. The aim of this study was to evaluate the associations between RDW and the clinical characteristics of pHCC patients. Medical records of 110 treatment-naive pHCC patients were retrospectively reviewed. Their clinical characteristics on admission, including RDW, liver function tests and tumor stage, were extracted, and their relationships were analyzed using Spearman correlation and Kruskal-Wallis test. Sixty-eight healthy individuals were set as controls. RDW was significantly increased in pHCC patients and correlated with the liver function tests. However, no correlation between RDW and tumor stage was found. RDW may be used to assess the liver function, but not the tumor stage in pHCC patients.

A smart core-sheath nanofiber that captures and releases red blood cells from the blood

NASA Astrophysics Data System (ADS)

Shi, Q.; Hou, J.; Zhao, C.; Xin, Z.; Jin, J.; Li, C.; Wong, S.-C.; Yin, J.

2016-01-01

A smart core-sheath nanofiber for non-adherent cell capture and release is demonstrated. The nanofibers are fabricated by single-spinneret electrospinning of poly(N-isopropylacrylamide) (PNIPAAm), polycaprolactone (PCL) and nattokinase (NK) solution blends. The self-assembly of PNIPAAm and PCL blends during the electrospinning generates the core-sheath PCL/PNIPAAm nanofibers with PNIPAAm as the sheath. The PNIPAAm-based core-sheath nanofibers are switchable between hydrophobicity and hydrophilicity with temperature change and enhance stability in the blood. When the nanofibers come in contact with blood, the NK is released from the nanofibers to resist platelet adhesion on the nanofiber surface, facilitating the direct capture and isolation of red blood cells (RBCs) from the blood above phase-transition temperature of PNIPAAm. Meanwhile, the captured RBCs are readily released from the nanofibers with temperature stimuli in an undamaged manner. The release efficiency of up to 100% is obtained while maintaining cellular integrity and function. This work presents promising nanofibers to effectively capture non-adherent cells and release for subsequent molecular analysis and diagnosis of single cells.A smart core-sheath nanofiber for non-adherent cell capture and release is demonstrated. The nanofibers are fabricated by single-spinneret electrospinning of poly(N-isopropylacrylamide) (PNIPAAm), polycaprolactone (PCL) and nattokinase (NK) solution blends. The self-assembly of PNIPAAm and PCL blends during the electrospinning generates the core-sheath PCL/PNIPAAm nanofibers with PNIPAAm as the sheath. The PNIPAAm-based core-sheath nanofibers are switchable between hydrophobicity and hydrophilicity with temperature change and enhance stability in the blood. When the nanofibers come in contact with blood, the NK is released from the nanofibers to resist platelet adhesion on the nanofiber surface, facilitating the direct capture and isolation of red blood cells (RBCs) from the blood above phase-transition temperature of PNIPAAm. Meanwhile, the captured RBCs are readily released from the nanofibers with temperature stimuli in an undamaged manner. The release efficiency of up to 100% is obtained while maintaining cellular integrity and function. This work presents promising nanofibers to effectively capture non-adherent cells and release for subsequent molecular analysis and diagnosis of single cells. Electronic supplementary information (ESI) available: Electrospinning of polymer nanofibers; FTIR spectra and XPS spectra of PCL, PNIPAAm and PCL/PNIPAAm nanofibers; SEM images of PCL/PNIPAAm nanofibers with varied composition; PNIPAAm content on the sheath of nanofibers; stability of core-sheath PCL/PNIPAAm nanofibers. Platelet adhesion on the PCL/PNIPAAm nanofibers in the presence of NK; Protein adsorption on nanofibers. See DOI: 10.1039/c5nr07070h

Profound re-organization of cell surface proteome in equine retinal pigment epithelial cells in response to in vitro culturing.

PubMed

Szober, Christoph M; Hauck, Stefanie M; Euler, Kerstin N; Fröhlich, Kristina J H; Alge-Priglinger, Claudia; Ueffing, Marius; Deeg, Cornelia A

2012-10-31

The purpose of this study was to characterize the cell surface proteome of native compared to cultured equine retinal pigment epithelium (RPE) cells. The RPE plays an essential role in visual function and represents the outer blood-retinal barrier. We are investigating immunopathomechanisms of equine recurrent uveitis, an autoimmune inflammatory disease in horses leading to breakdown of the outer blood-retinal barrier and influx of autoreactive T-cells into affected horses' vitrei. Cell surface proteins of native and cultured RPE cells from eye-healthy horses were captured by biotinylation, analyzed by high resolution mass spectrometry coupled to liquid chromatography (LC MS/MS), and the most interesting candidates were validated by PCR, immunoblotting and immunocytochemistry. A total of 112 proteins were identified, of which 84% were cell surface membrane proteins. Twenty-three of these proteins were concurrently expressed by both cell states, 28 proteins exclusively by native RPE cells. Among the latter were two RPE markers with highly specialized RPE functions: cellular retinaldehyde-binding protein (CRALBP) and retinal pigment epithelium-specific protein 65kDa (RPE65). Furthermore, 61 proteins were only expressed by cultured RPE cells and absent in native cells. As we believe that initiating events, leading to the breakdown of the outer blood-retinal barrier, take place at the cell surface of RPE cells as a particularly exposed barrier structure, this differential characterization of cell surface proteomes of native and cultured equine RPE cells is a prerequisite for future studies.

Endocrine pancreatic function changes after acute pancreatitis.

PubMed

Wu, Deqing; Xu, Yaping; Zeng, Yue; Wang, Xingpeng

2011-10-01

This study aimed to investigate the impairment of pancreatic endocrine function and the associated risk factors after acute pancreatitis (AP). Fifty-nine patients were subjected to tests of pancreatic function after an attack of pancreatitis. The mean time after the event was 3.5 years. Pancreatic endocrine function was evaluated by fasting blood glucose (FBG), glycosylated hemoglobin, fasting blood insulin, and C-peptide. Homeostasis model assessment was used to evaluate insulin resistance and islet β-cell function. Pancreatic exocrine function was evaluated by fecal elastase 1. Factors that could influence endocrine function were also investigated. Nineteen patients (32%) were found to have elevated FBG, whereas 5 (8%) had abnormal glycosylated hemoglobin levels. The levels of FBG, fasting blood insulin, and C-peptide were higher in patients than in controls (P < 0.01). The islet β-cell function of patients was lower than that of controls (P < 0.01), whereas insulin resistance index was higher among patients (P < 0.01). Obesity, hyperlipidemia, and diabetes-related symptoms were found to be associated with endocrine insufficiency. Pancreatic exocrine functional impairment was found at the same time. Endocrine functional impairment with insulin resistance was found in patients after AP. Obesity, hyperlipidemia, and diabetes-related symptoms increased the likelihood of developing functional impairment after AP.

The correlation between NK cell and liver function in patients with primary hepatocellular carcinoma.

PubMed

Sha, Wei Hong; Zeng, Xiao Hui; Min, Lu

2014-05-01

This study aimed to detect the expression of natural killer (NK) cell receptor natural killer group 2D (NKG2D) in the peripheral blood of patients with primary hepatocellular carcinoma and to discuss the correlation between NK cell cytotoxicity and liver function. The number of NK cells and the expression of NK cell receptor NKG2D in peripheral blood were determined by flow cytometry in patients with primary hepatocellular carcinoma, hepatitis B cirrhosis, chronic hepatitis B, and healthy controls. When compared with patients in the healthy and the chronic hepatitis B groups, the primary hepatocellular carcinoma group showed significant decreases in all parameters, including the cytotoxicity of NK cells on K562 cells, expression rate of NKG2D in NK cells, number of NKG2D(+) NK cells, expression level of NKG2D, and number of NK cells (p<0.05). The activity of NK cells showed a positive correlation, whereas the Child-Pugh scores in the primary hepatocellular carcinoma and the hepatitis B cirrhosis groups showed a negative correlation with all parameters detected above. The decrease of NK cell activity in patients with primary hepatocellular carcinoma is closely related to their lower expression of NKG2D. Liver function affects the expression of NKG2D and the activity of NK cells.

Increased adenosine triphosphate production by peripheral blood CD4+ cells in patients with hematologic malignancies treated with stem cell mobilization agents.

PubMed

Manga, Kiran; Serban, Geo; Schwartz, Joseph; Slotky, Ronit; Patel, Nita; Fan, Jianshe; Bai, Xiaolin; Chari, Ajai; Savage, David; Suciu-Foca, Nicole; Colovai, Adriana I

2010-07-01

Hematopoietic stem cell (HSC) transplantation is an important therapeutic option for patients with hematologic malignancies. To explore the immunomodulatory effects of HSC mobilization agents, we studied the function and phenotype of CD4(+) T cells from 16 adult patients with hematologic malignancies undergoing HSC mobilization treatment for autologous transplantation. Immune cell function was determined using the Immuknow (Cylex) assay by measuring the amount of adenosine triphosphate (ATP) produced by CD4(+) cells from whole blood. ATP activity measured in G-CSF-treated patients was significantly higher than that measured in healthy individuals or "nonmobilized" patients. In patients treated with G-CSF, CD4(+) T cells were predominantly CD25(low)FOXP3(low), consistent with an activated phenotype. However, T-cell depletion did not abrogate ATP production in blood samples from G-CSF-treated patients, indicating that CD4(+) myeloid cells contributed to the increased ATP levels observed in these patients. There was a significant correlation between ATP activity and patient survival, suggesting that efficient activation of CD4(+) cells during mobilization treatment predicts a low risk of disease relapse. Monitoring immune cell reactivity using the Immuknow assay may assist in the clinical management of patients with hematologic malignancies and optimization of HSC mobilization protocols. Copyright 2010 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.

STRESS-INDUCED REDISTRIBUTION OF IMMUNE CELLS - FROM BARRACKS TO BOULEVARDS TO BATTLEFIELDS: A TALE OF THREE HORMONES - CURT RICHTER AWARD WINNER

PubMed Central

Dhabhar, Firdaus S.; Malarkey, William B.; Neri, Eric; McEwen, Bruce S.

2012-01-01

Background The surveillance and effector functions of the immune system are critically dependent on the appropriate distribution of immune cells in the body. An acute or short-term stress response induces a rapid and significant redistribution of immune cells among different body compartments. Stress-induced leukocyte redistribution may be a fundamental survival response that directs leukocyte subpopulations to specific target organs during stress, and significantly enhances the speed, efficacy and regulation of an immune response. Immune responses are generally enhanced in compartments (e.g., skin) that are enriched with leukocytes, and suppressed in compartments that are depleted of leukocytes during/following stress. The experiments described here were designed to elucidate the: 1) Time-course, trajectory, and subpopulation-specificity of stress-induced mobilization and trafficking of blood leukocytes. 2) Individual and combined actions of the principal stress hormones, norepinephrine (NE), epinephrine (EPI), and corticosterone (CORT), in mediating mobilization or trafficking of specific leukocyte subpopulations. 3) Effects of stress/stress hormones on adhesion molecule, L-selectin (CD62L), expression by each subpopulation to assess its adhesion / functional / maturation status. Methods Male Sprague Dawley rats were stressed (short-term restraint, 2–120 min), or adrenalectomized and injected with vehicle (VEH), NE, EPI, CORT, or their combinations, and blood was collected for measurement of hormones and flow cytometric quantification of leukocyte subpopulations. Results Acute stress induced an early increase/mobilization of neutrophils, lymphocytes, helper T cells (Th), cytolytic T cells (CTL), and B cells into the blood, followed by a decrease/trafficking of all cell types out of the blood, except neutrophil numbers that continued to increase. CD62L expression was increased on neutrophils, decreased on Th, CTL, and natural killer (NK) cells, and showed a biphasic decrease on monocytes & B cells, suggesting that CD62L is involved in mediating the redistribution effects of stress. Additionally, we observed significant differences in the direction, magnitude, and subpopulation specificity of the effects of each hormone: NE increased leukocyte numbers, most notably CD62L−/+ neutrophils and CD62L− B cells. EPI increased monocyte and neutrophil numbers, most notably CD62L−/+ neutrophils and CD62L− monocytes, but decreased lymphocyte numbers with CD62L−/+ CTL and CD62L+ B cells being especially sensitive. CORT decreased monocyte, lymphocyte, Th, CTL, and B cell numbers with CD62L− and CD62L+ cells being equally affected. Thus, naïve (CD62L+) vs. memory (CD62L−) T cells, classical (CD62L+) vs. non-classical (CD62L−) monocytes, and similarly distinct functional subsets of other leukocyte populations are differentially mobilized into the blood and trafficked to tissues by stress hormones. Conclusion Stress hormones orchestrate a large-scale redistribution of immune cells in the body. NE and EPI mobilize immune cells into the bloodstream, and EPI and CORT induce traffic out of the blood possibly to tissue surveillance pathways, lymphoid tissues, and sites of ongoing or de novo immune activation. Immune cell subpopulations appear to show differential sensitivities and redistribution responses to each hormone depending on the type of leukocyte (neutrophil, monocyte or lymphocyte) and its maturation/functional characteristics (e.g., resident or inflammatory monocyte, naïve or central/effector memory T cell). Thus, stress hormones could be administered simultaneously or sequentially to induce specific leukocyte subpopulations to be mobilized into the blood, or to traffic from blood to tissues. Stress hormone-mediated changes in immune cell distribution could be clinically harnessed to: 1) Direct leukocytes to sites of vaccination, wound healing, infection, or cancer and thereby enhance protective immunity. 2) Reduce leukocyte traffic to sites of inflammatory/autoimmune reactions. 3) Sequester immune cells in relatively protected compartments to minimize exposure to cytotoxic treatments like radiation or localized chemotherapy. 4) Measure biological resistance/sensitivity to stress hormones in vivo. In keeping with the guidelines for Richter Award manuscripts, in addition to original data we also present a model and synthesis of findings in the context of the literature on the effects of short-term stress on immune cell distribution and function. PMID:22727761

Modeled Microgravity Inhibits Apoptosis in Peripheral Blood Lymphocytes

NASA Technical Reports Server (NTRS)

Risin, Diana; Pellis, Neal R.

2000-01-01

Microgravity interferes with numerous lymphocyte functions (expression of cell surface molecules, locomotion, polyclonal and antigen-specific activation, and the protein kinase C activity in signal transduction). The latter suggests that gravity may also affect programmed cell death (PCD) in lymphocyte populations. To test this hypothesis, we investigated spontaneous, activation- and radiation-induced PCD in peripheral blood mononuclear cells (PBMC) exposed to modeled microgravity using a rotating cell culture system. The results showed significant inhibition of radiation- and activation-induced apoptosis in modeled microgravity and provide insights into the potential mechanisms of this phenomenon.

Modeled microgravity inhibits apoptosis in peripheral blood lymphocytes

NASA Technical Reports Server (NTRS)

Risin, D.; Pellis, N. R.; McIntire, L. V. (Principal Investigator)

2001-01-01

Microgravity interferes with numerous lymphocyte functions (expression of cell surface molecules, locomotion, polyclonal and antigen-specific activation, and the protein kinase C activity in signal transduction). The latter suggests that gravity may also affect programmed cell death (PCD) in lymphocyte populations. To test this hypothesis, we investigated spontaneous, activation- and radiation-induced PCD in peripheral blood mononuclear cells exposed to modeled microgravity (MMG) using a rotating cell culture system. The results showed significant inhibition of radiation- and activation-induced apoptosis in MMG and provide insights into the potential mechanisms of this phenomenon.

Analyzing Structure and Function of Vascularization in Engineered Bone Tissue by Video-Rate Intravital Microscopy and 3D Image Processing.

PubMed

Pang, Yonggang; Tsigkou, Olga; Spencer, Joel A; Lin, Charles P; Neville, Craig; Grottkau, Brian

2015-10-01

Vascularization is a key challenge in tissue engineering. Three-dimensional structure and microcirculation are two fundamental parameters for evaluating vascularization. Microscopic techniques with cellular level resolution, fast continuous observation, and robust 3D postimage processing are essential for evaluation, but have not been applied previously because of technical difficulties. In this study, we report novel video-rate confocal microscopy and 3D postimage processing techniques to accomplish this goal. In an immune-deficient mouse model, vascularized bone tissue was successfully engineered using human bone marrow mesenchymal stem cells (hMSCs) and human umbilical vein endothelial cells (HUVECs) in a poly (D,L-lactide-co-glycolide) (PLGA) scaffold. Video-rate (30 FPS) intravital confocal microscopy was applied in vitro and in vivo to visualize the vascular structure in the engineered bone and the microcirculation of the blood cells. Postimage processing was applied to perform 3D image reconstruction, by analyzing microvascular networks and calculating blood cell viscosity. The 3D volume reconstructed images show that the hMSCs served as pericytes stabilizing the microvascular network formed by HUVECs. Using orthogonal imaging reconstruction and transparency adjustment, both the vessel structure and blood cells within the vessel lumen were visualized. Network length, network intersections, and intersection densities were successfully computed using our custom-developed software. Viscosity analysis of the blood cells provided functional evaluation of the microcirculation. These results show that by 8 weeks, the blood vessels in peripheral areas function quite similarly to the host vessels. However, the viscosity drops about fourfold where it is only 0.8 mm away from the host. In summary, we developed novel techniques combining intravital microscopy and 3D image processing to analyze the vascularization in engineered bone. These techniques have broad applicability for evaluating vascularization in other engineered tissues as well.

Myeloid-derived suppressor cells can be efficiently generated from human hematopoietic progenitors and peripheral blood monocytes.

PubMed

Casacuberta-Serra, Sílvia; Parés, Marta; Golbano, Arantxa; Coves, Elisabet; Espejo, Carmen; Barquinero, Jordi

2017-07-01

Myeloid-derived suppressor cells (MDSCs) have an important role in controlling inflammation. As such, they are both a therapeutic target and, based on the administration of ex vivo-generated MDSCs, a therapeutic tool. However, there are relatively few reports describing methods to generate human MDSCs, and most of them rely on cells obtained from peripheral blood monocytes. We investigated alternative approaches to the generation of MDSCs from hematopoietic progenitors and monocytes. Purified CD34 + hematopoietic progenitors from apheresis products and CD14 + cells isolated from buffy coats were cultured in the presence of different combinations of cytokines. The resulting myeloid cell populations were then characterized phenotypically and functionally. Progenitor cells cultured in the presence of SCF+TPO+FLT3-L+GM-CSF+IL-6 gave rise to both monocytic (M)- and granulocytic (G)-MDSCs but production of the latter was partially inhibited by IL-3. M-MDSCs but not G-MDSCs were obtained by culturing peripheral blood monocytes with GM-CSF+IL-6 or GM-CSF+TGF-β1 for 6 days. CD14 expression was downregulated in the cultured cells. PD-L1 expression at baseline was lower in hematopoietic progenitor cell-derived than in monocyte-derived MDSCs, but was markedly increased in response to stimulation with LPS+IFN-γ. The functionality of the two MDSC subtypes was confirmed in studies of the suppression of allogeneic and mitogen-induced proliferation and by cytokine profiling. Here we describe both the culture conditions that allow the generation of MDSCs and the phenotypical and functional characterization of these cell populations.

NMDA Receptors Regulate Genes Responsible for Major Immune Functions of Mononuclears in Human Peripheral Blood.

PubMed

Kuzmina, U Sh; Zainullina, L F; Sadovnikov, S V; Vakhitov, V A; Vakhitova, Yu V

2018-06-19

To determine the role of NMDA receptors in the functional regulation of immunocompetent cells, comparative assay was carried out for genes expressed in the mononuclears in peripheral blood of healthy persons under normal conditions and after blockade of these receptors. The genes, whose expression changed in response to blockade of NMDA receptors in mononuclears, encode the products involved in regulation of the major functions of immune cells, such as proliferation (IL4, VCAM1, and CDKN2A), apoptosis (BAX, MYC, CDKN2A, HSPB1, and CADD45A), activation (IL4R, IL4, VCAM1, and CDKN2A), and differentiation (IL4, VCAM1, and BAX).

How hormones influence composition and physiological function of the brain-blood barrier.

PubMed

Hampl, R; Bičíková, M; Sosvorová, L

2015-01-01

Hormones exert many actions in the brain. Their access and effects in the brain are regulated by the blood-brain barrier (BBB). Hormones as other substances may enter the brain and vice versa either by paracellular way requiring breaching tight junctions stitching the endothelial cells composing the BBB, or by passage through the cells (transcellular way). Hormones influence both ways through their receptors, both membrane and intracellular, present on/in the BBB. In the review the main examples are outlined how hormones influence the expression and function of proteins forming the tight junctions, as well as how they regulate expression and function of major protein transporters mediating transport of various substances including hormone themselves.

A Synopsis of Factors Regulating Beta Cell Development and Beta Cell Mass

PubMed Central

Prasadan, Krishna; Shiota, Chiyo; Xiangwei, Xiao; Ricks, David; Fusco, Joseph; Gittes, George

2016-01-01

The insulin-secreting beta cells in the endocrine pancreas regulate blood glucose levels, and loss of functional beta cells leads to insulin deficiency, hyperglycemia (high blood glucose) and diabetes mellitus. Current treatment strategies for type-1 (autoimmune) diabetes are islet transplantation, which has significant risks and limitations, or normalization of blood glucose with insulin injections, which is clearly not ideal. The type-1 patients can lack insulin counter-regulatory mechanism; therefore, hypoglycemia is a potential risk. Hence, a cell-based therapy offers a better alternative for the treatment of diabetes. Past research was focused on attempting to generate replacement beta cells from stem cells, however, recently there has been an increasing interest in identifying mechanisms that will lead to the conversion of pre-existing differentiated endocrine cells into beta cells. The goal of this review is to provide an overview of several of the key factors that regulate new beta cell formation (neogenesis) and beta cell proliferation. PMID:27105622

Renal intercalated cells and blood pressure regulation.

PubMed

Wall, Susan M

2017-12-01

Type B and non-A, non-B intercalated cells are found within the connecting tubule and the cortical collecting duct. Of these cell types, type B intercalated cells are known to mediate Cl - absorption and HCO 3 - secretion largely through pendrin-dependent Cl - /HCO 3 - exchange. This exchange is stimulated by angiotensin II administration and is also stimulated in models of metabolic alkalosis, for instance after aldosterone or NaHCO 3 administration. In some rodent models, pendrin-mediated HCO 3 - secretion modulates acid-base balance. However, the role of pendrin in blood pressure regulation is likely of more physiological or clinical significance. Pendrin regulates blood pressure not only by mediating aldosterone-sensitive Cl - absorption, but also by modulating the aldosterone response for epithelial Na + channel (ENaC)-mediated Na + absorption. Pendrin regulates ENaC through changes in open channel of probability, channel surface density, and channels subunit total protein abundance. Thus, aldosterone stimulates ENaC activity through both direct and indirect effects, the latter occurring through its stimulation of pendrin expression and function. Therefore, pendrin contributes to the aldosterone pressor response. Pendrin may also modulate blood pressure in part through its action in the adrenal medulla, where it modulates the release of catecholamines, or through an indirect effect on vascular contractile force. This review describes how aldosterone and angiotensin II-induced signaling regulate pendrin and the contributory role of pendrin in distal nephron function and blood pressure.

Hydrogen inhalation ameliorated mast cell-mediated brain injury after intracerebral hemorrhage in mice.

PubMed

Manaenko, Anatol; Lekic, Tim; Ma, Qingyi; Zhang, John H; Tang, Jiping

2013-05-01

Hydrogen inhalation was neuroprotective in several brain injury models. Its mechanisms are believed to be related to antioxidative stress. We investigated the potential neurovascular protective effect of hydrogen inhalation especially effect on mast cell activation in a mouse model of intracerebral hemorrhage. Controlled in vivo laboratory study. Animal research laboratory. One hundred seventy-one 8-week-old male CD-1 mice were used. Collagenase-induced intracerebral hemorrhage model in 8-week-old male CD-1 mice was used. Hydrogen was administrated via spontaneous inhalation. The blood-brain barrier permeability and neurologic deficits were investigated at 24 and 72 hours after intracerebral hemorrhage. Mast cell activation was evaluated by Western blot and immuno-staining. The effects of hydrogen inhalation on mast cell activation were confirmed in an autologous blood injection model intracerebral hemorrhage. At 24 and 72 hours post intracerebral hemorrhage, animals showed blood-brain barrier disruption, brain edema, and neurologic deficits, accompanied with phosphorylation of Lyn kinase and release of tryptase, indicating mast cell activation. Hydrogen treatment diminished phosphorylation of Lyn kinase and release of tryptase, decreased accumulation and degranulation of mast cells, attenuated blood-brain barrier disruption, and improved neurobehavioral function. Activation of mast cells following intracerebral hemorrhage contributed to increase of blood-brain barrier permeability and brain edema. Hydrogen inhalation preserved blood-brain barrier disruption by prevention of mast cell activation after intracerebral hemorrhage.

A close look at brain dynamics: cells and vessels seen by in vivo two-photon microscopy.

PubMed

Fumagalli, Stefano; Ortolano, Fabrizio; De Simoni, Maria-Grazia

2014-10-01

The cerebral vasculature has a unique role in providing a constant supply of oxygen and nutrients to ensure normal brain functions. Blood vessels that feed the brain are far from being simply channels for passive transportation of fluids. They form complex structures made up of different cell types. These structures regulate blood supply, local concentrations of O2 and CO2, transport of small molecules, trafficking of plasma cells and fine cerebral functions in normal and diseased brains. Until few years ago, analysis of these functions has been typically based on post mortem techniques, whose interpretation is limited by the need for tissue processing at specific times. For a reliable and effective picture of the dynamic processes in the central nervous system, real-time information in vivo is required. There are now few in vivo systems, among which two-photon microscopy (2-PM) is a truly innovative tool for studying the brain. 2-PM has been used to dissect specific aspects of vascular and immune cell dynamics in the context of neurological diseases, providing exciting results that could not have been obtained with conventional methods. This review summarizes the latest findings on vascular and immune system action in the brain, with particular focus on the dynamic responses after ischemic brain injury. 2-PM has helped define the hierarchical architecture of the brain vasculature, the dynamic interaction between the vasculature and immune cells recruited to lesion sites, the effects of blood flow on neuronal and microglial activity and the ability of cells of the neurovascular unit to regulate blood flow. Copyright © 2014 Elsevier Ltd. All rights reserved.

Change in permeability of the plasma membrane of blood cells in irradiated animals

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shevchenko, A.S.; Kobyalko, V.O.; Lazarev, N.M.

1994-11-01

The Chernobyl nuclear disaster showed the exposure of the thyroid gland to radioactive iodine is an important factor of radiation damage to animals. Examination of domestic animals showed a marked inhibition of thyroid hormone secretion and changes in red cell membrane permeability for calcium in the absence of marked hematological shifts. At the same time the disturbed thyroid statis is associated with changes in some structural and functional parameters of blood cells. This research on calves shows that radiation damage to the thyroid produces a modifying effect on blood cell membrane permeability for calcium during both the acute and latemore » periods following exposure to 131I. 15 refs., 2 figs., 1 tab.« less

Micro-array isolation of circulating tumor cells (CTCs): the droplet biopsy chip

NASA Astrophysics Data System (ADS)

Panchapakesan, B.

2017-08-01

We present a new method for circulating tumor cell capture based on micro-array isolation from droplets. Called droplet biopsy, our technique uses a 76-element array of carbon nanotube devices functionalized with anti-EpCAM and antiHer2 antibodies for immunocapture of spiked breast cancer cells in the blood. This droplet biopsy chip can enable capture of CTCs based on both positive and negative selection strategy. Negative selection is achieved through depletion of contaminating leukocytes through the differential settling of blood into layers. We report 55%-100% cancer cell capture yield in this first droplet biopsy chip study. The droplet biopsy is an enabling idea where one can capture CTCs based on multiple biomarkers in a single blood sample.

Hematological shift in goat kids naturally devoid of prion protein.

PubMed

Reiten, Malin R; Bakkebø, Maren K; Brun-Hansen, Hege; Lewandowska-Sabat, Anna M; Olsaker, Ingrid; Tranulis, Michael A; Espenes, Arild; Boysen, Preben

2015-01-01

The physiological role of the cellular prion protein (PrP(C)) is incompletely understood. The expression of PrP(C) in hematopoietic stem cells and immune cells suggests a role in the development of these cells, and in PrP(C) knockout animals altered immune cell proliferation and phagocytic function have been observed. Recently, a spontaneous nonsense mutation at codon 32 in the PRNP gene in goats of the Norwegian Dairy breed was discovered, rendering homozygous animals devoid of PrP(C). Here we report hematological and immunological analyses of homozygous goat kids lacking PrP(C) (PRNP(Ter/Ter) ) compared to heterozygous (PRNP (+/Ter)) and normal (PRNP (+/+)) kids. Levels of cell surface PrP(C) and PRNP mRNA in peripheral blood mononuclear cells (PBMCs) correlated well and were very low in PRNP (Ter/Ter), intermediate in PRNP (+/Ter) and high in PRNP (+/+) kids. The PRNP (Ter/Ter) animals had a shift in blood cell composition with an elevated number of red blood cells (RBCs) and a tendency toward a smaller mean RBC volume (P = 0.08) and an increased number of neutrophils (P = 0.068), all values within the reference ranges. Morphological investigations of blood smears and bone marrow imprints did not reveal irregularities. Studies of relative composition of PBMCs, phagocytic ability of monocytes and T-cell proliferation revealed no significant differences between the genotypes. Our data suggest that PrP(C) has a role in bone marrow physiology and warrant further studies of PrP(C) in erythroid and immune cell progenitors as well as differentiated effector cells also under stressful conditions. Altogether, this genetically unmanipulated PrP(C)-free animal model represents a unique opportunity to unveil the enigmatic physiology and function of PrP(C).

Th17 Cells Coordinate with Th22 Cells in Maintaining Homeostasis of Intestinal Tissues and both are Depleted in SIV-Infected Macaques.

PubMed

Xu, Huanbin; Wang, Xiaolei; Veazey, Ronald S

2014-05-01

Th17 and Th22 cells are thought to function as innate regulators of mucosal antimicrobial responses, tissue inflammation and mucosal integrity, yet their role in persistent SIV infection is still unclear. Here we compared Th17 and Th22 cells in their phenotype, effector/cytokine function, and frequency in blood and intestinal mucosal tissues, and correlate levels with mucosal damage in SIV-infected rhesus macaques. We found that Th17/Th22 cells share similar features in that both highly produce TNF-α and IL-2 and express CCR5 in intestinal tissues; yet very few show cytotoxic functions, as evidenced by lack of IFN-γ and granzyme B production. Further, Th17/Th22 cells display distinct tissue-specific distributions. Both Th17 and Th22 cells and cytokine secretion were significantly depleted in both blood and intestine in chronically SIV-infected macaques. The frequency of Th17 and Th22 cells in the intestine positively correlated with percentages of intestinal CD4+ T cells and negatively with damage to intestinal mucosa, and plasma viral loads in SIV infection. These findings indicate Th17 and Th22 cells share considerable functions, and may coordinate in innate mucosal immune responses, and their regional loss in the intestine may be associated with local mucosal immune dysfunction in persistent HIV/SIV infection.

Th17 Cells Coordinate with Th22 Cells in Maintaining Homeostasis of Intestinal Tissues and both are Depleted in SIV-Infected Macaques

PubMed Central

Xu, Huanbin; Wang, Xiaolei; Veazey, Ronald S.

2014-01-01

Th17 and Th22 cells are thought to function as innate regulators of mucosal antimicrobial responses, tissue inflammation and mucosal integrity, yet their role in persistent SIV infection is still unclear. Here we compared Th17 and Th22 cells in their phenotype, effector/cytokine function, and frequency in blood and intestinal mucosal tissues, and correlate levels with mucosal damage in SIV-infected rhesus macaques. We found that Th17/Th22 cells share similar features in that both highly produce TNF-α and IL-2 and express CCR5 in intestinal tissues; yet very few show cytotoxic functions, as evidenced by lack of IFN-γ and granzyme B production. Further, Th17/Th22 cells display distinct tissue-specific distributions. Both Th17 and Th22 cells and cytokine secretion were significantly depleted in both blood and intestine in chronically SIV-infected macaques. The frequency of Th17 and Th22 cells in the intestine positively correlated with percentages of intestinal CD4+ T cells and negatively with damage to intestinal mucosa, and plasma viral loads in SIV infection. These findings indicate Th17 and Th22 cells share considerable functions, and may coordinate in innate mucosal immune responses, and their regional loss in the intestine may be associated with local mucosal immune dysfunction in persistent HIV/SIV infection. PMID:25364618

Tilted pillar array fabrication by the combination of proton beam writing and soft lithography for microfluidic cell capture: Part 1 Design and feasibility.

PubMed

Rajta, Istvan; Huszánk, Robert; Szabó, Atilla T T; Nagy, Gyula U L; Szilasi, Szabolcs; Fürjes, Peter; Holczer, Eszter; Fekete, Zoltan; Járvás, Gabor; Szigeti, Marton; Hajba, Laszlo; Bodnár, Judit; Guttman, Andras

2016-02-01

Design, fabrication, integration, and feasibility test results of a novel microfluidic cell capture device is presented, exploiting the advantages of proton beam writing to make lithographic irradiations under multiple target tilting angles and UV lithography to easily reproduce large area structures. A cell capture device is demonstrated with a unique doubly tilted micropillar array design for cell manipulation in microfluidic applications. Tilting the pillars increased their functional surface, therefore, enhanced fluidic interaction when special bioaffinity coating was used, and improved fluid dynamic behavior regarding cell culture injection. The proposed microstructures were capable to support adequate distribution of body fluids, such as blood, spinal fluid, etc., between the inlet and outlet of the microfluidic sample reservoirs, offering advanced cell capture capability on the functionalized surfaces. The hydrodynamic characteristics of the microfluidic systems were tested with yeast cells (similar size as red blood cells) for efficient capture. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Log (TG)/HDL-C ratio as a predictor of decreased islet beta cell function in patients with type 2 diabetes: 6-year cohort study.

PubMed

Zhou, Meicen; Li, Zengyi; Min, Rui; Dong, Yaxiu; Sun, Qi; Li, Yuxiu

2015-09-01

The aim of the present study was to explore whether the triglyceride to high density lipoprotein cholesterol ratio [log (TG)/HDL-C] and peripheral blood leukocytes DNA telomere length could predict future islet beta cell function decreased in Chinese type 2 diabetes mellitus (T2DM) during a 6-year cohort. Sixty T2DM patients (without insulin treatment at baseline) were included in the 6-year cohort study. Peripheral blood leukocytes DNA telomere length, HbA1c, blood lipid profile, fatty fat acid, glucose, insulin and C peptide (3 h after a mixed meal) were determined. Delta C peptide area under curve (Delta CP AUC) was used to reflect change in beta cell secretion function (Delta CP AUC = baseline CP AUC - CP AUC after 6 years). Subjects were divided into slow decrease of beta cell function group (Delta CP AUCslow group) and fast decrease group (Delta CP AUCfast group) according to median of Delta CP AUC. Baseline demographic characteristics, clinical variables between two groups were compared. Correlations between baseline data and Delta CP AUC were analyzed. Baseline log (TG)/HDL-C was positively correlated with Delta CP AUC (r = 0.306, P = 0.027); log (TG)/HDL-C in Delta CP AUCfast group was higher than that in Delta CP AUCslow group (0.103 ± 0.033 vs 0.083 ± 0.030, P = 0.027). There was no significant difference in DNA telomere length between the two groups. Change in DNA telomere length over 6 years was not significantly correlated with baseline blood lipid. In Chinese T2DM patients, high baseline log (TG)/HDL-C ratio predicts fast progression of islet beta cell dysfunction. It may be a simple index to predict progression speed of islet beta cell dysfunction. © 2014 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd.

HIV Infection in Uncircumcised Men Is Associated With Altered CD8 T-cell Function But Normal CD4 T-cell Numbers in the Foreskin.

PubMed

Prodger, Jessica L; Hirbod, Taha; Gray, Ronald; Kigozi, Godfrey; Nalugoda, Fred; Galiwango, Ronald; Reynolds, Steven J; Huibner, Sanja; Wawer, Maria J; Serwadda, David; Kaul, Rupert

2014-04-15

Human immunodeficiency virus (HIV)-infected (HIV+) men are more susceptible to sexually transmitted infections, and may be superinfected by HIV. We hypothesized that HIV induces immune alterations in the foreskin that may impact the subsequent acquisition/clearance of genital coinfections. Foreskin tissue and blood were obtained from 70 HIV-uninfected and 20 HIV+ men undergoing circumcision. T cells were characterized by flow cytometry, immunohistochemistry, and polymerase chain reaction. There was substantial influx of CD8 T-cells into the foreskins of HIV+ men (108.8 vs 23.1 cells/mm(2); P < .001); but foreskin CD4 T-cell density was unchanged (43.0 vs 33.7/mm(2); P = .67), despite substantial blood depletion (409.0 vs 877.8 cells/µL; P < .001). While frequencies of foreskin C-C chemokine receptor type 5(+) (CCR5(+)) T cells, T regulatory cells, and T-helper 17 cells were unaltered in HIV+ men, CD8 T-cell production of tumor necrosis factor α (TNFα) was decreased. HIV-specific CD8 T cells were present in the foreskins of HIV+ men, although their frequency and function was reduced compared to the blood. Foreskin CD4 T-cell density and CCR5 expression were not reduced during HIV infection, perhaps explaining susceptibility to HIV superinfection. Foreskin CD8 T-cell density was increased, but decreased production of TNFα may enhance susceptibility to genital coinfections in HIV+ men.

New findings on venous thrombogenesis

PubMed Central

Byrnes, James R.; Wolberg, Alisa S.

2017-01-01

Summary Venous thrombosis (VT) is the third most common cause of cardiovascular death worldwide. Complications from VT and pulmonary embolism are the leading cause of lost disability-adjusted life years. Risks include genetic (e.g., non-O blood group, activated protein C resistance, hyperprothrombinemia) and acquired (e.g., age, surgery, cancer, pregnancy, immobilisation, female hormone use) factors. Pathophysiologic mechanisms that promote VT are incompletely understood, but involve abnormalities in blood coagulability, vessel function, and flow (so-called Virchow’s Triad). Epidemiologic studies of humans, animal models, and biochemical and biophysical investigations have revealed contributions from extrinsic, intrinsic, and common pathways of coagulation, endothelial cells, leukocytes, red blood cells, platelets, cell-derived microvesicles, stasis-induced changes in vascular cells, and blood rheology. Knowledge of these mechanisms may yield new therapeutic targets. Characterisation of mechanisms that mediate VT formation and stability, particularly in aging, are needed to advance understanding of VT. PMID:27878206

Tight junction protein expression and barrier properties of immortalized mouse brain microvessel endothelial cells.

PubMed

Brown, Rachel C; Morris, Andrew P; O'Neil, Roger G

2007-01-26

Understanding the molecular and biochemical mechanisms regulating the blood-brain barrier is aided by in vitro model systems. Many studies have used primary cultures of brain microvessel endothelial cells for this purpose. However, primary cultures limit the generation of material for molecular and biochemical assays since cells grow slowly, are prone to contamination by other neurovascular unit cells, and lose blood-brain barrier characteristics when passaged. To address these issues, immortalized cell lines have been generated. In these studies, we assessed the suitability of the immortalized mouse brain endothelial cell line, bEnd3, as a blood-brain barrier model. RT-PCR and immunofluorescence indicated expression of multiple tight junction proteins. bEnd3 cells formed barriers to radiolabeled sucrose, and responded like primary cultures to disrupting stimuli. Exposing cells to serum-free media on their basolateral side significantly decreased paracellular permeability; astrocyte-conditioned media did not enhance barrier properties. The serum-free media-induced decrease in permeability was correlated with an increase in claudin-5 and zonula occludens-1 immunofluorescence at cell-cell contracts. We conclude that bEnd3 cells are an attractive candidate as a model of the blood-brain barrier due to their rapid growth, maintenance of blood-brain barrier characteristics over repeated passages, formation of functional barriers and amenability to numerous molecular interventions.

TIGHT JUNCTION PROTEIN EXPRESSION AND BARRIER PROPERTIES OF IMMORTALIZED MOUSE BRAIN MICROVESSEL ENDOTHELIAL CELLS

PubMed Central

Brown, Rachel C.; Morris, Andrew P.; O’Neil, Roger G.

2007-01-01

Understanding the molecular and biochemical mechanisms regulating the blood-brain barrier is aided by in vitro model systems. Many studies have used primary cultures of brain microvessel endothelial cells for this purpose. However, primary cultures limit the generation of material for molecular and biochemical assays since cells grow slowly, are prone to contamination by other neurovascular unit cells, and lose blood-brain barrier characteristics when passaged. To address these issues, immortalized cell lines have been generated. In these studies, we assessed the suitability of the immortalized mouse brain endothelial cell line, bEnd3, as a blood-brain barrier model. RT-PCR and immunofluorescence indicated expression of multiple tight junction proteins. bEnd3 cells formed barriers to radiolabeled sucrose, and responded like primary cultures to disrupting stimuli. Exposing cells to serum-free media on their basolateral side significantly decreased paracellular permeability; astrocyte-conditioned media did not enhance barrier properties. The serum-free media-induced decrease in permeability was correlated with an increase in claudin-5 and zonula occludens-1 immunofluorescence at cell-cell contracts. We conclude that bEnd3 cells are an attractive candidate as a model of the blood-brain barrier due to their rapid growth, maintenance of blood-brain barrier characteristics over repeated passages, formation of functional barriers and amenability to numerous molecular interventions. PMID:17169347

Single-cell measurement of red blood cell oxygen affinity.

PubMed

Di Caprio, Giuseppe; Stokes, Chris; Higgins, John M; Schonbrun, Ethan

2015-08-11

Oxygen is transported throughout the body by hemoglobin (Hb) in red blood cells (RBCs). Although the oxygen affinity of blood is well-understood and routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of RBC volume and Hb concentration are taken millions of times per day by clinical hematology analyzers, and they are important factors in determining the health of the hematologic system. To better understand the variability and determinants of oxygen affinity on a cellular level, we have developed a system that quantifies the oxygen saturation, cell volume, and Hb concentration for individual RBCs in high throughput. We find that the variability in single-cell saturation peaks at an oxygen partial pressure of 2.9%, which corresponds to the maximum slope of the oxygen-Hb dissociation curve. In addition, single-cell oxygen affinity is positively correlated with Hb concentration but independent of osmolarity, which suggests variation in the Hb to 2,3-diphosphoglycerate (2-3 DPG) ratio on a cellular level. By quantifying the functional behavior of a cellular population, our system adds a dimension to blood cell analysis and other measurements of single-cell variability.

Single-cell measurement of red blood cell oxygen affinity

PubMed Central

Di Caprio, Giuseppe; Stokes, Chris; Higgins, John M.; Schonbrun, Ethan

2015-01-01

Oxygen is transported throughout the body by hemoglobin (Hb) in red blood cells (RBCs). Although the oxygen affinity of blood is well-understood and routinely assessed in patients by pulse oximetry, variability at the single-cell level has not been previously measured. In contrast, single-cell measurements of RBC volume and Hb concentration are taken millions of times per day by clinical hematology analyzers, and they are important factors in determining the health of the hematologic system. To better understand the variability and determinants of oxygen affinity on a cellular level, we have developed a system that quantifies the oxygen saturation, cell volume, and Hb concentration for individual RBCs in high throughput. We find that the variability in single-cell saturation peaks at an oxygen partial pressure of 2.9%, which corresponds to the maximum slope of the oxygen–Hb dissociation curve. In addition, single-cell oxygen affinity is positively correlated with Hb concentration but independent of osmolarity, which suggests variation in the Hb to 2,3-diphosphoglycerate (2–3 DPG) ratio on a cellular level. By quantifying the functional behavior of a cellular population, our system adds a dimension to blood cell analysis and other measurements of single-cell variability. PMID:26216973

Meningeal mast cells affect early T cell central nervous system infiltration and blood-brain barrier integrity through TNF: a role for neutrophil recruitment?

PubMed

Sayed, Blayne A; Christy, Alison L; Walker, Margaret E; Brown, Melissa A

2010-06-15

Mast cells contribute to the pathogenesis of experimental autoimmune encephalomyelitis, a rodent model of the human demyelinating disease multiple sclerosis. Yet their site and mode of action is unknown. In both diseases, myelin-specific T cells are initially activated in peripheral lymphoid organs. However, for disease to occur, these cells must enter the immunologically privileged CNS through a breach in the relatively impermeable blood-brain barrier. In this study, we demonstrate that a dense population of resident mast cells in the meninges, structures surrounding the brain and spinal cord, regulate basal CNS barrier function, facilitating initial T cell CNS entry. Through the expression of TNF, mast cells recruit an early wave of neutrophils to the CNS. We propose that neutrophils in turn promote the blood-brain barrier breach and together with T cells lead to further inflammatory cell influx and myelin damage. These findings provide specific targets for intervention in multiple sclerosis as well as other immune-mediated CNS diseases.

The Asymmetric Cell Division Regulators Par3, Scribble and Pins/Gpsm2 Are Not Essential for Erythroid Development or Enucleation

PubMed Central

Wölwer, Christina B.; Gödde, Nathan; Pase, Luke B.; Elsum, Imogen A.; Lim, Krystle Y. B.; Sacirbegovic, Faruk; Walkley, Carl R.; Ellis, Sarah; Ohno, Shigeo; Matsuzaki, Fumio; Russell, Sarah M.; Humbert, Patrick O.

2017-01-01

Erythroid enucleation is the process by which the future red blood cell disposes of its nucleus prior to entering the blood stream. This key event during red blood cell development has been likened to an asymmetric cell division (ACD), by which the enucleating erythroblast divides into two very different daughter cells of alternate molecular composition, a nucleated cell that will be removed by associated macrophages, and the reticulocyte that will mature to the definitive erythrocyte. Here we investigated gene expression of members of the Par, Scribble and Pins/Gpsm2 asymmetric cell division complexes in erythroid cells, and functionally tested their role in erythroid enucleation in vivo and ex vivo. Despite their roles in regulating ACD in other contexts, we found that these polarity regulators are not essential for erythroid enucleation, nor for erythroid development in vivo. Together our results put into question a role for cell polarity and asymmetric cell division in erythroid enucleation. PMID:28095473

Cancer cells remodel themselves and vasculature to overcome the endothelial barrier.

PubMed

Shenoy, Anitha K; Lu, Jianrong

2016-10-01

Metastasis refers to the spread of cancer cells from a primary tumor to distant organs mostly via the bloodstream. During the metastatic process, cancer cells invade blood vessels to enter circulation, and later exit the vasculature at a distant site. Endothelial cells that line blood vessels normally serve as a barrier to the movement of cells into or out of the blood. It is thus critical to understand how metastatic cancer cells overcome the endothelial barrier. Epithelial cancer cells acquire increased motility and invasiveness through epithelial-to-mesenchymal transition (EMT), which enables them to move toward vasculature. Cancer cells also express a variety of adhesion molecules that allow them to attach to vascular endothelium. Finally, cancer cells secrete or induce growth factors and cytokines to actively prompt vascular hyperpermeability that compromises endothelial barrier function and facilitates transmigration of cancer cells through the vascular wall. Elucidation of the mechanisms underlying metastatic dissemination may help develop new anti-metastasis therapeutics. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Lysosomal storage diseases and the blood-brain barrier.

PubMed

Begley, David J; Pontikis, Charles C; Scarpa, Maurizio

2008-01-01

The blood-brain barrier becomes a crucial issue in neuronopathic lysosomal storage diseases for three reasons. Firstly, the function of the blood-brain barrier may be compromised in many of the lysosomal storage diseases and this barrier dysfunction may contribute to the neuropathology seen in the diseases and accelerate cell death. Secondly, the substrate reduction therapies, which successfully reduce peripheral lysosomal storage, because of the blood-brain barrier may not have as free an access to brain cells as they do to peripheral cells. And thirdly, enzyme replacement therapy appears to have little access to the central nervous system as the mannose and mannose-6-phosphate receptors involved in their cellular uptake and transport to the lysosome do not appear to be expressed at the adult blood-brain barrier. This review will discuss in detail these issues and their context in the development of new therapeutic strategies.

Comparison of hindlimb unloading and partial weight suspension models for spaceflight-type condition induced effects on white blood cells

NASA Astrophysics Data System (ADS)

Wilson, Jolaine M.; Krigsfeld, Gabriel S.; Sanzari, Jenine K.; Wagner, Erika B.; Mick, Rosemarie; Kennedy, Ann R.

2012-01-01

Animal models are frequently used to assist in the determination of the long- and short-term effects of space flight. The space environment, including microgravity, can impact many physiological and immunological system parameters. It has been found that ground based models of microgravity produce changes in white blood cell counts, which negatively affects immunologic function. As part of the Center of Acute Radiation Research (CARR), we compared the acute effects on white blood cell parameters induced by the more traditionally used animal model of hindlimb unloading (HU) with a recently developed reduced weightbearing analog known as partial weight suspension (PWS). Female ICR mice were either hindlimb unloaded or placed in the PWS system at 16% quadrupedal weightbearing for 4 h, 1, 2, 7 or 10 days, at which point complete blood counts were obtained. Control animals (jacketed and non-jacketed) were exposed to identical conditions without reduced weightbearing. Results indicate that significant changes in total white blood cell (WBC), neutrophil, lymphocyte, monocyte and eosinophil counts were observed within the first 2 days of exposure to each system. These differences in blood cell counts normalized by day 7 in both systems. The results of these studies indicate that there are some statistically significant changes observed in the blood cell counts for animals exposed to both the PWS and HU simulated microgravity systems.

Glycolysis-mediated control of blood-brain barrier development and function.

PubMed

Salmina, Alla B; Kuvacheva, Natalia V; Morgun, Andrey V; Komleva, Yulia K; Pozhilenkova, Elena A; Lopatina, Olga L; Gorina, Yana V; Taranushenko, Tatyana E; Petrova, Lyudmila L

2015-07-01

The blood-brain barrier (BBB) consists of differentiated cells integrating in one ensemble to control transport processes between the central nervous system (CNS) and peripheral blood. Molecular organization of BBB affects the extracellular content and cell metabolism in the CNS. Developmental aspects of BBB attract much attention in recent years, and barriergenesis is currently recognized as a very important and complex mechanism of CNS development and maturation. Metabolic control of angiogenesis/barriergenesis may be provided by glucose utilization within the neurovascular unit (NVU). The role of glycolysis in the brain has been reconsidered recently, and it is recognized now not only as a process active in hypoxic conditions, but also as a mechanism affecting signal transduction, synaptic activity, and brain development. There is growing evidence that glycolysis-derived metabolites, particularly, lactate, affect barriergenesis and functioning of BBB. In the brain, lactate produced in astrocytes or endothelial cells can be transported to the extracellular space via monocarboxylate transporters (MCTs), and may act on the adjoining cells via specific lactate receptors. Astrocytes are one of the major sources of lactate production in the brain and significantly contribute to the regulation of BBB development and functioning. Active glycolysis in astrocytes is required for effective support of neuronal activity and angiogenesis, while endothelial cells regulate bioavailability of lactate for brain cells adjusting its bidirectional transport through the BBB. In this article, we review the current knowledge with regard to energy production in endothelial and astroglial cells within the NVU. In addition, we describe lactate-driven mechanisms and action of alternative products of glucose metabolism affecting BBB structural and functional integrity in developing and mature brain. Copyright © 2015 Elsevier Ltd. All rights reserved.

Anti-leukemia activity of a bacterial toxin with natural specificity for LFA-1 on white blood cells

PubMed Central

Kachlany, Scott C.; Schwartz, Amy B.; Balashova, Nataliya V.; Hioe, Catarina E.; Tuen, Michael; Le, Amy; Kaur, Manpreet; Mei, Yongyi; Rao, Jia

2009-01-01

The oral bacterium, Aggregatibacter actinomycetemcomitans, produces a leukotoxin (LtxA) that is specific for white blood cells (WBCs) from humans and Old World primates by interacting with lymphocyte function antigen-1 (LFA-1) on susceptible cells. To determine if LtxA could be used as a therapeutic agent for the treatment of WBC diseases, we tested the in vitro and in vivo anti-leukemia activity of the toxin. LtxA kills human malignant WBC lines and primary leukemia cells from acute myeloid leukemia patients, but healthy peripheral blood mononuclear cells (PBMCs) are relatively resistant to LtxA-mediated cytotoxicity. Levels of LFA-1 on cell lines correlated with killing by LtxA and the toxin preferentially killed cells expressing the activated form of LFA-1. In a SCID mouse model for human leukemia, LtxA had potent therapeutic value resulting in long-term survival in LtxA-treated mice. Intravenous infusion of LtxA into a rhesus macaque resulted in a drop in WBC counts at early times post-infusion; however, red blood cells, platelets, hemoglobin and blood chemistry values remained unaffected. Thus, LtxA may be an effective and safe novel therapeutic agent for the treatment of hematologic malignancies. PMID:19747730

Breaking the Blood-Brain Barrier With Mannitol to Aid Stem Cell Therapeutics in the Chronic Stroke Brain.

PubMed

Tajiri, Naoki; Lee, Jea Young; Acosta, Sandra; Sanberg, Paul R; Borlongan, Cesar V

2016-01-01

Blood-brain barrier (BBB) permeabilizers, such as mannitol, can facilitate peripherally delivered stem cells to exert therapeutic benefits on the stroke brain. Although this BBB permeation-aided stem cell therapy has been demonstrated in the acute stage of stroke, such BBB permeation in the chronic stage of the disease remains to be examined. Adult Sprague-Dawley rats initially received sham surgery or experimental stroke via the 1-h middle cerebral artery occlusion (MCAo) model. At 1 month after the MCAo surgery, stroke animals were randomly assigned to receive human umbilical cord stem cells only (2 million viable cells), mannitol only (1.1 mol/L mannitol at 4°C), combined human umbilical cord stem cells (200,000 viable cells) and mannitol (1.1 mol/L mannitol at 4°C), and vehicle (phosphate-buffered saline) only. Stroke animals that received human umbilical cord blood cells alone or combined human umbilical cord stem cells and mannitol exhibited significantly improved motor performance and significantly better brain cell survival in the peri-infarct area compared to stroke animals that received vehicle or mannitol alone, with mannitol treatment reducing the stem cell dose necessary to afford functional outcomes. Enhanced neurogenesis in the subventricular zone accompanied the combined treatment of human umbilical cord stem cells and mannitol. We showed that BBB permeation facilitates the therapeutic effects of a low dose of peripherally transplanted stem cells to effectively cause functional improvement and increase neurogenesis in chronic stroke.

Solanum Nigrum polysaccharide (SNL) extract effects in transplanted tumor-bearing mice--erythrocyte membrane fluidity and blocking of functions.

PubMed

Yuan, Hong-Liang; Liu, Xiao-Lei; Liu, Ying-Jie

2014-01-01

Solanum nigrum L. has been used in traditional Chinese medicine because of its diuretic and antipyretic effects. The present research concerned effects of crude polysaccharides isolated from Solanum nigrum L. on erythrocyte membranes of tumor-bearing S180 and H22 in mice. Fluorescence- labeled red blood cell membranes were used with DPH fluorescence spectrophotometry to examine erythrocyte membrane fluidity, and colorimetry to determine degree of erythrocyte surface membrane blocking. Extent of reaction by tumor-bearing mice with the enzyme erythrocyte membrane bubble shadow detection of red cell membrane variation in the degree of closure before and after administration. Solanum nigrum polysaccharide could significantly improve the S180 and H22 tumor-bearing mice erythrocyte membrane fluidity, compared with the control group, the difference was significant (p<0.01), SNL can significantly improve the red blood cell membrane and then S180 tumor-bearing mice sealing ability, compared with the negative control group, the difference was significant(p<0.05, p<0.01). H22 tumor-bearing mice can increase red cell membrane and then sealing ability, the difference was significant (p<0.05). Solanum nigrum polysaccharide degree of fluidity and blocking two transplanted tumors in mice restored the ability to raise the red cell membrane has a significant effect. Solanum nigrum L.-type mice transplanted tumor can affect the red blood cell membrane fluidity and re-closed, through the red cell membrane of red blood cells to enhance the immune function of the possibility of erythrocyte immunity against tumor formation garland provide experimental basis.

Proliferative Potentials of Bone Marrow and Blood Cells Studied by in vitro Uptake of H 3-Thymidine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bond, V. P.; Fliedner, T. M.; Cronkite, E. P.

1959-01-01

Cell proliferative activity and potential in the circulating blood and in the bone marrow of individuals with normal hematopoiesis, and in patients with hematopoietic dyscrasias was studied by means of in vitro one hour incubation with tritiated thymidine (H 3Th) and 6tripping film autoradiography. The labeled material is incorporated only into DNA during synthesis. In normal bone marrow, labeling was seen at 1 hour in all cell lineages, and in cells variously referred to as "reticulum," "stem,'' " stroma,'' etc., cells. Erythropoietic cells were labeled as far as the polychromatic normoblast; the myeloid series was labeled to the myelocyte state.more » Leukemia cells in the bone marrow and peripheral blood of patients with acute or chronic myelocytic leukemia incorporated label avidly; the small typical leukemia cell of chronic lymphocytic leukemia did not label at all. Less than 3 per cent of the myeloma cells in patients with multiple myeloma incorporated thymidine. Most striking was the finding of small numbers of labeled large mononuclear cells of different morphological types in the peripheral blood of normal human beings, and an increase in the number of morphologically identical cells in the blood of patients with infection and infectious mononucleosis. The labeling indicates active DNA synthesis and thus these cells presumably are capable of division. It is suggested that these cells may represent a mobile pool of primitive progenitor cells and are multipotential in their function.« less

[Allogenic hematopoietic stem cell transplantation with unrelated cord blood: report of three cases from the Chilean cord blood bank].

PubMed

Barriga, Francisco; Wietstruck, Angélica; Rojas, Nicolás; Bertin, Pablo; Pizarro, Isabel; Carmona, Amanda; Guilof, Alejandro; Rojas, Iván; Oyarzún, Enrique

2013-08-01

Public cord blood banks are a source of hematopoietic stem cells for patients with hematological diseases who lack a family donor and need allogeneic transplantation. In June 2007 we started a cord blood bank with units donated in three maternity wards in Santiago, Chile. We report the first three transplants done with cord blood units form this bank. Cord blood units were obtained by intrauterine collection at delivery. They were depleted of plasma and red cells and frozen in liquid nitrogen. Tests for total nucleated cells, CD34 cell content, viral serology, bacterial cultures and HLA A, B and DRB1 were done. Six hundred cord blood units were stored by March 2012. Three patients received allogeneic transplant with cord blood from our bank, two with high risk lymphoblastic leukemia and one with severe congenital anemia. They received conditioning regimens according to their disease and usual supportive care for unrelated donor transplantation until full hematopoietic and immune reconstitution was achieved. The three patients had early engraftment of neutrophils and platelets. The child corrected his anemia and the leukemia patients remain in complete remission. The post-transplant course was complicated with Epstein Barr virus, cytomegalovirus and BK virus infection. Two patients are fully functional 24 and 33 months after transplant, the third is still receiving immunosuppression.

Tumor Necrosis Factor-producing T-regulatory Cells Are Associated With Severe Liver Injury in Patients With Acute Hepatitis A.

PubMed

Choi, Yoon Seok; Jung, Min Kyung; Lee, Jeewon; Choi, Seong Jin; Choi, Sung Hoon; Lee, Hyun Woong; Lee, Jong-Joo; Kim, Hyung Joon; Ahn, Sang Hoon; Lee, Dong Hyeon; Kim, Won; Park, Su-Hyung; Huh, Jun R; Kim, Hyoung-Pyo; Park, Jun Yong; Shin, Eui-Cheol

2018-03-01

CD4 + CD25 + Foxp3 + T-regulatory (Treg) cells control immune responses and maintain immune homeostasis. However, under inflammatory conditions, Treg cells produce cytokines that promote inflammation. We investigated production of tumor necrosis factor (TNF) by Treg cells in patients with acute hepatitis A (AHA), and examined the characteristics of these cells and association with clinical factors. We analyzed blood samples collected from 63 patients with AHA at the time of hospitalization (and some at later time points) and 19 healthy donors in South Korea. Liver tissues were collected from patients with fulminant AHA during liver transplantation. Peripheral blood mononuclear cells were isolated from whole blood and lymphocytes were isolated from liver tissues and analyzed by flow cytometry. Cytokine production from Treg cells (CD4 + CD25 + Foxp3 + ) was measured by immunofluorescence levels following stimulation with anti-CD3 and anti-CD28. Epigenetic stability of Treg cells was determined based on DNA methylation patterns. Phenotypes of Treg cells were analyzed by flow cytometry and an RORγt inhibitor, ML-209, was used to inhibit TNF production. Treg cell suppression assay was performed by co-culture of Treg-depleted peripheral blood mononuclear cells s and isolated Treg cells. A higher proportion of CD4 + CD25 + Foxp3 + Treg cells from patients with AHA compared with controls produced TNF upon stimulation with anti-CD3 and anti-CD28 (11.2% vs 2.8%). DNA methylation analysis confirmed the identity of the Treg cells. TNF-producing Treg cells had features of T-helper 17 cells, including up-regulation of RORγt, which was required for TNF production. The Treg cells had reduced suppressive functions compared with Treg cells from controls. The frequency of TNF-producing Treg cells in AHA patients' blood correlated with their serum level of alanine aminotransferase. Treg cells from patients with AHA have altered functions compared with Treg cells from healthy individuals. Treg cells from patients with AHA produce higher levels of TNF, gain features of T-helper 17 cells, and have reduced suppressive activity. The presence of these cells is associated with severe liver injury in patients with AHA. Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.

2-D Model for Normal and Sickle Cell Blood Microcirculation

NASA Astrophysics Data System (ADS)

Tekleab, Yonatan; Harris, Wesley

2011-11-01

Sickle cell disease (SCD) is a genetic disorder that alters the red blood cell (RBC) structure and function such that hemoglobin (Hb) cannot effectively bind and release oxygen. Previous computational models have been designed to study the microcirculation for insight into blood disorders such as SCD. Our novel 2-D computational model represents a fast, time efficient method developed to analyze flow dynamics, O2 diffusion, and cell deformation in the microcirculation. The model uses a finite difference, Crank-Nicholson scheme to compute the flow and O2 concentration, and the level set computational method to advect the RBC membrane on a staggered grid. Several sets of initial and boundary conditions were tested. Simulation data indicate a few parameters to be significant in the perturbation of the blood flow and O2 concentration profiles. Specifically, the Hill coefficient, arterial O2 partial pressure, O2 partial pressure at 50% Hb saturation, and cell membrane stiffness are significant factors. Results were found to be consistent with those of Le Floch [2010] and Secomb [2006].

Red blood cells inhibit activation-induced cell death and oxidative stress in human peripheral blood T lymphocytes.

PubMed

Fonseca, A M; Porto, G; Uchida, K; Arosa, F A

2001-05-15

Red blood cells (RBCs) are known to perform one prominent function: to carry and deliver oxygen to the tissues. Earlier studies, however, suggested a role for RBCs in potentiating T-cell proliferation in vitro. Here it is shown that the presence of RBCs in cultures of stimulated human peripheral blood lymphocytes strengthens T-cell proliferation and survival. Analysis of phosphatidylserine externalization and DNA fragmentation showed that RBCs inhibit T-cell apoptosis. This inhibition correlated with a reduction in CD71 but not CD95 expression. RBCs enhanced T-cell proliferation and survival upon activation with phytohemagglutinin and with OKT3 antibodies. Studies aimed at characterizing the cellular and molecular basis of the protection afforded to T cells by RBCs showed that (1) optimal protection required intact RBCs and red cell/T-cell contact but not monocytes; (2) RBCs markedly reduced the level of intracellular reactive oxygen species; and (3) RBCs inhibited the formation of protein-bound acrolein, a peroxidation adduct in biologic systems. Overall, these data indicate that human RBCs protect T cells from activation-induced cell death, at least in part by reducing the pro-oxidant state, and suggest a role for RBCs as conceivable modulators of T-cell homeostasis.

Cerebrovascular function and cognition in childhood: a systematic review of transcranial doppler studies

PubMed Central

2014-01-01

Background The contribution of cerebrovascular function to cognitive performance is gaining increased attention. Transcranial doppler (TCD) is portable, reliable, inexpensive and extremely well tolerated by young and clinical samples. It enables measurement of blood flow velocity in major cerebral arteries at rest and during cognitive tasks. Methods We systematically reviewed evidence for associations between cognitive performance and cerebrovascular function in children (0-18 years), as measured using TCD. A total of 2778 articles were retrieved from PsychInfo, Pubmed, and EMBASE searches and 25 relevant articles were identified. Results Most studies investigated clinical groups, where decreased blood flow velocities in infants were associated with poor neurological functioning, and increased blood flow velocities in children with Sickle cell disease were typically associated with cognitive impairment and lower intelligence. Studies were also identified assessing autistic behaviour, mental retardation and sleep disordered breathing. In healthy children, the majority of studies reported cognitive processing produced lateralised changes in blood flow velocities however these physiological responses did not appear to correlate with behavioural cognitive performance. Conclusion Poor cognitive performance appears to be associated with decreased blood flow velocities in premature infants, and increased velocities in Sickle cell disease children using TCD methods. However knowledge in healthy samples is relatively limited. The technique is well tolerated by children, is portable and inexpensive. It therefore stands to make a valuable contribution to knowledge regarding the underlying functional biology of cognitive performance in childhood. PMID:24602446

Does oral supplementation of a fermented papaya preparation correct respiratory burst function of innate immune cells in type 2 diabetes mellitus patients?

PubMed

Dickerson, Ryan; Banerjee, Jaideep; Rauckhorst, Adam; Pfeiffer, Douglas R; Gordillo, Gayle M; Khanna, Savita; Osei, Kwame; Roy, Sashwati

2015-02-01

Fermented papaya preparation (FPP) is a nutritional supplement reported to act as an antioxidant by scavenging reactive oxygen species (ROS) and removing "bad ROS," while inducing "respiratory burst" production of necessary "good ROS." We sought to investigate the safety of oral administration of FPP (9 g/day, 6 weeks) to T2D patients with regard to its effect on the hyperglycemia status of these patients. Peripheral blood was collected during a baseline visit, followed by subsequent collections both during and after supplementation. Induced "respiratory burst" ROS production was measured at each visit in addition to fasting blood glucose, lipid profile, glycated hemoglobin (HbA1c), and lipid/protein peroxidation. Oral FPP supplementation induced "respiratory burst" in peripheral blood mononuclear cells while not influencing other blood parameters studied. When human monocytic THP-1 cells were supplemented with sugar-based FPP, cellular ATP and NADPH concentrations were increased while matched glucose alone did not produce similar effects, suggesting a glucose-independent component of FPP to be responsible for increasing cellular energetics. THP-1 cells supplemented with FPP also exhibited higher mitochondrial membrane potential (Δψm) and oxygen consumption as compared with cells treated with glucose alone. Taken together, our observations lead to the hypothesis that FPP corrects inducible "respiratory burst" function in type 2 diabetes patients.

Circulating Blood eNOS Contributes to the Regulation of Systemic Blood Pressure and Nitrite Homeostasis

PubMed Central

Wood, Katherine C.; Cortese-Krott, Miriam M.; Kovacic, Jason C.; Noguchi, Audrey; Liu, Virginia B.; Wang, Xunde; Raghavachari, Nalini; Boehm, Manfred; Kato, Gregory J.; Kelm, Malte; Gladwin, Mark T.

2013-01-01

Objective Mice genetically deficient in endothelial nitric oxide synthase (eNOS−/−) are hypertensive with lower circulating nitrite levels, indicating the importance of constitutively produced nitric oxide (NO•) to blood pressure regulation and vascular homeostasis. While the current paradigm holds that this bioactivity derives specifically from expression of eNOS in endothelium, circulating blood cells also express eNOS protein. A functional red cell eNOS that modulates vascular NO• signaling has been proposed. Approach and Results To test the hypothesis that blood cells contribute to mammalian blood pressure regulation via eNOS-dependent NO• generation, we cross-transplanted WT and eNOS−/− mice, producing chimeras competent or deficient for eNOS expression in circulating blood cells. Surprisingly, we observed a significant contribution of both endothelial and circulating blood cell eNOS to blood pressure and systemic nitrite levels, the latter being a major component of the circulating NO• reservoir. These effects were abolished by the NOS inhibitor L-NAME and repristinated by the NOS substrate L-Arginine, and were independent of platelet or leukocyte depletion. Mouse erythrocytes were also found to carry an eNOS protein and convert 14C-Arginine into 14C-Citrulline in a NOS-dependent fashion. Conclusions These are the first studies to definitively establish a role for a blood borne eNOS, using cross transplant chimera models, that contributes to the regulation of blood pressure and nitrite homeostasis. This work provides evidence suggesting that erythrocyte eNOS may mediate this effect. PMID:23702660

Mesenchymal Stem Cell/Red Blood Cell-Inspired Nanoparticle Therapy in Mice with Carbon Tetrachloride-Induced Acute Liver Failure.

PubMed

Liang, Hongxia; Huang, Ke; Su, Teng; Li, Zhenhua; Hu, Shiqi; Dinh, Phuong-Uyen; Wrona, Emily A; Shao, Chen; Qiao, Li; Vandergriff, Adam C; Hensley, M Taylor; Cores, Jhon; Allen, Tyler; Zhang, Hongyu; Zeng, Qinglei; Xing, Jiyuan; Freytes, Donald O; Shen, Deliang; Yu, Zujiang; Cheng, Ke

2018-06-26

Acute liver failure is a critical condition characterized by global hepatocyte death and often time needs a liver transplantation. Such treatment is largely limited by donor organ shortage. Stem cell therapy offers a promising option to patients with acute liver failure. Yet, therapeutic efficacy and feasibility are hindered by delivery route and storage instability of live cell products. We fabricated a nanoparticle that carries the beneficial regenerative factors from mesenchymal stem cells and further coated it with the membranes of red blood cells to increase blood stability. Unlike uncoated nanoparticles, these particles promote liver cell proliferation in vitro and have lower internalization by macrophage cells. After intravenous delivery, these artificial stem cell analogs are able to remain in the liver and mitigate carbon tetrachloride-induced liver failure in a mouse model, as gauged by histology and liver function test. Our technology provides an innovative and off-the-shelf strategy to treat liver failure.

Concise Review: Therapeutic Potential of Adipose Tissue-Derived Angiogenic Cells

PubMed Central

Brinchmann, Jan E.

2012-01-01

Inadequate blood supply to tissues is a leading cause of morbidity and mortality today. Ischemic symptoms caused by obstruction of arterioles and capillaries are currently not treatable by vessel replacement or dilatation procedures. Therapeutic angiogenesis, the treatment of tissue ischemia by promoting the proliferation of new blood vessels, has recently emerged as one of the most promising therapies. Neovascularization is most often attempted by introduction of angiogenic cells from different sources. Emerging evidence suggests that adipose tissue (AT) is an excellent reservoir of autologous cells with angiogenic potential. AT yields two cell populations of importance for neovascularization: AT-derived mesenchymal stromal cells, which likely act predominantly as pericytes, and AT-derived endothelial cells (ECs). In this concise review we discuss different physiological aspects of neovascularization, briefly present cells isolated from the blood and bone marrow with EC properties, and then discuss isolation and cell culture strategies, phenotype, functional capabilities, and possible therapeutic applications of angiogenic cells obtained from AT. PMID:23197872

Autologous blood cell therapies from pluripotent stem cells

PubMed Central

Lengerke, Claudia; Daley, George Q.

2010-01-01

Summary The discovery of human embryonic stem cells (hESCs) raised promises for a universal resource for cell based therapies in regenerative medicine. Recently, fast-paced progress has been made towards the generation of pluripotent stem cells (PSCs) amenable for clinical applications, culminating in reprogramming of adult somatic cells to autologous PSCs that can be indefinitely expanded in vitro. However, besides the efficient generation of bona fide, clinically safe PSCs (e.g. without the use of oncoproteins and gene transfer based on viruses inserting randomly into the genome), a major challenge in the field remains how to efficiently differentiate PSCs to specific lineages and how to select for cells that will function normally upon transplantation in adults. In this review, we analyse the in vitro differentiation potential of PSCs to the hematopoietic lineage discussing blood cell types that can be currently obtained, limitations in derivation of adult-type HSCs and prospects for clinical application of PSCs-derived blood cells. PMID:19910091

Density-Gradient Mediated Band Extraction of Leukocytes from Whole Blood Using Centrifugo-Pneumatic Siphon Valving on Centrifugal Microfluidic Discs

PubMed Central

Kearney, Sinéad M.; Kilcawley, Niamh A.; Early, Philip L.; Glynn, Macdara T.; Ducrée, Jens

2016-01-01

Here we present retrieval of Peripheral Blood Mononuclear Cells by density-gradient medium based centrifugation for subsequent analysis of the leukocytes on an integrated microfluidic “Lab-on-a-Disc” cartridge. Isolation of white blood cells constitutes a critical sample preparation step for many bioassays. Centrifugo-pneumatic siphon valves are particularly suited for blood processing as they function without need of surface treatment and are ‘low-pass’, i.e., holding at high centrifugation speeds and opening upon reduction of the spin rate. Both ‘hydrostatically’ and ‘hydrodynamically’ triggered centrifugo-pneumatic siphon valving schemes are presented. Firstly, the geometry of the pneumatic chamber of hydrostatically primed centrifugo-pneumatic siphon valves is optimised to enable smooth and uniform layering of blood on top of the density-gradient medium; this feature proves to be key for efficient Peripheral Blood Mononuclear Cell extraction. A theoretical analysis of hydrostatically primed valves is also presented which determines the optimum priming pressure for the individual valves. Next, ‘dual siphon’ configurations for both hydrostatically and hydrodynamically primed centrifugo-pneumatic siphon valves are introduced; here plasma and Peripheral Blood Mononuclear Cells are extracted through a distinct siphon valve. This work represents a first step towards enabling on disc multi-parameter analysis. Finally, the efficiency of Peripheral Blood Mononuclear Cells extraction in these structures is characterised using a simplified design. A microfluidic mechanism, which we termed phase switching, is identified which affects the efficiency of Peripheral Blood Mononuclear Cell extraction. PMID:27167376

Functional dilatation and medial remodeling of the renal artery in response to chronic increased blood flow.

PubMed

Roan, Jun-Neng; Yeh, Chin-Yi; Chiu, Wen-Cheng; Lee, Chou-Hwei; Chang, Shih-Wei; Jiangshieh, Ya-Fen; Tsai, Yu-Chuan; Lam, Chen-Fuh

2011-01-01

Renal blood flow (RBF) is tightly regulated by several intrinsic pathways in maintaining optimal kidney blood supply. Using a rat model of aortocaval (AC) fistula, we investigated remodeling of the renal artery following prolonged increased blood flow. An AC fistula was created in the infrarenal aorta of anesthetized rats, and changes of blood flow in the renal artery were assessed using an ultrasonic flow probe. Morphological changes and expression of endothelial nitric oxide synthase and matrix metalloproteinase-2 in the remodeled renal artery were analyzed. Blood flow in the renal artery increased immediately after creation of AC fistula, but normal RBF was restored 8 weeks later. The renal artery dilated significantly 8 weeks after operation. Expression of endothelial nitric oxide synthase and matrix metalloproteinase-2 was upregulated shortly after blood flow increase, and returned to baseline levels after 3 weeks. Histological sections showed luminal dilatation with medial thickening and endothelial cell-to-smooth muscle cell attachments in the remodeled renal artery. Increased RBF was accommodated by functional dilatation and remodeling in the medial layer of the renal artery in order to restore normal blood flow. Our results provide important mechanistic insight into the intrinsic regulation of the renal artery in response to increased RBF. Copyright © 2011 S. Karger AG, Basel.

Characterization of γδ regulatory T cells from peripheral blood in patients with multiple myeloma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ma, Yongyong; Department of Hematology, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, 325000; Lei, Huyi

γδ regulatory T cells are able to inhibit the activation and function of T cells involved in antigen-specific immune responses. This study aimed to investigate the potential role of γδ regulatory T cells in inhibiting anti-tumor immune responses in patients diagnosed as multiple myeloma (MM). We measured the levels of γδ T cells, the distribution and clonally amplified TCR Vγ and VδT cells in peripheral blood of healthy donors, patients recently diagnosed with MM, and MM patients in remission cohorts. In addition, we evaluated the ability of γδ regulatory T cells to inhibit the proliferation of CD4+CD25- T cells andmore » detected the expression of immunoregulatory-associated molecules. We found that the levels of γδ regulatory T cells from the peripheral blood in patients of MM were significantly higher than those in healthy donors. Comparison of γδT regulatory cells function in MM and healthy donors showed similarly inhibitory effects on the proliferation of T cells. Additionally, TLR8 expression level increased significantly in MM patients compared to healthy donors, while the expression levels of Foxp3, CD25, CTLA4, GITR, GATA3 and Tbet in MM patients and healthy donors showed no significant difference. Taken together, our study reveals the potential role of γδ regulatory T cells in inhibiting anti-tumor immune responses in MM patients.« less

Stem cell transplantation (cord blood transplants).

PubMed

Chao, Nelson J; Emerson, Stephen G; Weinberg, Kenneth I

2004-01-01

Allogeneic stem cell transplantation is an accepted treatment modality for selected malignant and non-malignant diseases. However, the ability to identify suitably matched related or unrelated donors can be difficult in some patients. Alternative sources of stem cells such as cord blood provide a readily available graft for such patients. Data accumulated over the past several years have demonstrated that the use of cord blood is an accepted source of stem cells for pediatric patients. Since the cell numbers of hematopoietic progenitors in cord blood is limited and the collection can occur only in a single occasion, its use in adult patients can be more problematic. Here, new developments in the use of cord blood for adults and studies aimed at expansion of cord blood cells and immune reconstitution are described. In Section I, Dr. Nelson Chao describes the early data in cord blood transplantation in adult patients. The patient outcomes are reviewed and analyzed for various factors such as cell dose, HLA typing, and patient selection that could have contributed to the final outcome of these adult patients. Myeloablative as well as nonmyeloablative approaches are presented. Discussion of the various benefits and risks are presented. More recent data from multiple single institutions as well as larger registry data comparisons are also provided. Analyses of these studies suggest methods to improve on the outcome. These newer data should lead to a logical progression in the use of cord blood cells in adult patients. In Section II, Dr. Stephen Emerson describes the historical efforts associated with expansion of hematopoietic stem cells, specifically with cord blood cells. These efforts to expand cord blood cells continue with novel methods. Moreover, a better understanding of stem cell biology and signaling is critical if we are to be able to effectively expand these cells for clinical use. An alternative, more direct, approach to expanding stem cells could be achieved by specific genetic pathways known or believed to support primitive HSC proliferation such as Notch-1 receptor activation, Wnt/LEF-1 pathway induction, telomerase or the Homeobox (Hox) gene products. The clinical experience with the use of expanded cord blood cells is also discussed. In Section III, Dr. Kenneth Weinberg describes immune reconstitution or lack thereof following cord blood transplantation. One of the hallmarks of successful hematopoietic stem cell transplantation is the ability to fully reconstitute the immune system of the recipient. Thus, the relationship between stem cell source and the development of T lymphocyte functions required for protection of the recipient from infection will be described, and cord blood recipients will be compared with those receiving other sources of stem cells. T cell development is described in detail, tracking from prethymic to postthymic lymphocytes with specific attention to umbilical cord blood as the source of stem cells. Moreover, a discussion of the placenta as a special microenvironment for umbilical cord blood is presented. Strategies to overcome the immunological defects are presented to improve the outcome of these recipients.

Human induced pluripotent stem cells can reach complete terminal maturation: in vivo and in vitro evidence in the erythropoietic differentiation model

PubMed Central

Kobari, Ladan; Yates, Frank; Oudrhiri, Noufissa; Francina, Alain; Kiger, Laurent; Mazurier, Christelle; Rouzbeh, Shaghayegh; El-Nemer, Wassim; Hebert, Nicolas; Giarratana, Marie-Catherine; François, Sabine; Chapel, Alain; Lapillonne, Hélène; Luton, Dominique; Bennaceur-Griscelli, Annelise; Douay, Luc

2012-01-01

Background Human induced pluripotent stem cells offer perspectives for cell therapy and research models for diseases. We applied this approach to the normal and pathological erythroid differentiation model by establishing induced pluripotent stem cells from normal and homozygous sickle cell disease donors. Design and Methods We addressed the question as to whether these cells can reach complete erythroid terminal maturation notably with a complete switch from fetal to adult hemoglobin. Sickle cell disease induced pluripotent stem cells were differentiated in vitro into red blood cells and characterized for their terminal maturation in terms of hemoglobin content, oxygen transport capacity, deformability, sickling and adherence. Nucleated erythroblast populations generated from normal and pathological induced pluripotent stem cells were then injected into non-obese diabetic severe combined immunodeficiency mice to follow the in vivo hemoglobin maturation. Results We observed that in vitro erythroid differentiation results in predominance of fetal hemoglobin which rescues the functionality of red blood cells in the pathological model of sickle cell disease. We observed, in vivo, the switch from fetal to adult hemoglobin after infusion of nucleated erythroid precursors derived from either normal or pathological induced pluripotent stem cells into mice. Conclusions These results demonstrate that human induced pluripotent stem cells: i) can achieve complete terminal erythroid maturation, in vitro in terms of nucleus expulsion and in vivo in terms of hemoglobin maturation; and ii) open the way to generation of functionally corrected red blood cells from sickle cell disease induced pluripotent stem cells, without any genetic modification or drug treatment. PMID:22733021

Induction of suppressor cells from peripheral blood T cells by 15-hydroperoxyeicosatetraenoic acid (15-HPETE).

PubMed

Gualde, N; Rigaud, M; Goodwin, J S

1985-11-01

15-hydroperoxyeicosetetraenoic acid (15-HPETE), a lipoxygenase metabolite of arachidonic acid, inhibited polyclonal IgG and IgM production in pokeweed mitogen (PWM)-stimulated cultures of human peripheral blood mononuclear cells, whereas 15-hydroxyeicosetetraenoic acid (15-HETE) had little effect in this system. T cells preincubated for 18 hr with 15-HPETE caused substantial inhibition of IgG and IgM production of fresh, autologous B and T cells stimulated by PWM. The suppressive effect of the 15-HPETE-treated cells was lost if the cells were irradiated before the PWM culture, but not by treatment with mitomycin C. The suppressive effect was also lost if OKT8+ T cells were removed after, but not before, preincubation of the T cells with 15-HPETE. OKT8- T cells incubated with 15-HPETE for 18 hr showed a large increase in the percentage of cells staining with directly fluoresceinated Leu-2, another marker for suppressor cells. Thus, 15-HPETE induces functional and phenotypic suppressor cells from resting human peripheral blood T cells.

Isolation of Human Skin Dendritic Cell Subsets.

PubMed

Gunawan, Merry; Jardine, Laura; Haniffa, Muzlifah

2016-01-01

Dendritic cells (DCs) are specialized leukocytes with antigen-processing and antigen-presenting functions. DCs can be divided into distinct subsets by anatomical location, phenotype and function. In human, the two most accessible tissues to study leukocytes are peripheral blood and skin. DCs are rare in human peripheral blood (<1 % of mononuclear cells) and have a less mature phenotype than their tissue counterparts (MacDonald et al., Blood. 100:4512-4520, 2002; Haniffa et al., Immunity 37:60-73, 2012). In contrast, the skin covering an average total surface area of 1.8 m(2) has approximately tenfold more DCs than the average 5 L of total blood volume (Wang et al., J Invest Dermatol 134:965-974, 2014). DCs migrate spontaneously from skin explants cultured ex vivo, which provide an easy method of cell isolation (Larsen et al., J Exp Med 172:1483-1493, 1990; Lenz et al., J Clin Invest 92:2587-2596, 1993; Nestle et al., J Immunol 151:6535-6545, 1993). These factors led to the extensive use of skin DCs as the "prototype" migratory DCs in human studies. In this chapter, we detail the protocols to isolate DCs and resident macrophages from human skin. We also provide a multiparameter flow cytometry gating strategy to identify human skin DCs and to distinguish them from macrophages.

Biology of the blood-nerve barrier and its alteration in immune mediated neuropathies.

PubMed

Kanda, Takashi

2013-02-01

The blood-nerve barrier (BNB) is a dynamic and competent interface between the endoneurial microenvironment and the surrounding extracellular space or blood. It is localised at the innermost layer of the multilayered ensheathing perineurium and endoneurial microvessels, and is the key structure that controls the internal milieu of the peripheral nerve parenchyma. Since the endoneurial BNB is the point of entry for pathogenic T cells and various soluble factors, including cytokines, chemokines and immunoglobulins, understanding this structure is important to prevent and treat human immune mediated neuropathies such as Guillain-Barré syndrome, chronic inflammatory demyelinating polyneuropathy, POEMS (polyneuropathy, organomegaly, endocrinopathy, monoclonal protein and skin changes) syndrome and a subset of diabetic neuropathy. However, compared with the blood-brain barrier, only limited knowledge has been accumulated regarding the function, cell biology and clinical significance of the BNB. This review describes the basic structure and functions of the endoneurial BNB, provides an update of the biology of the cells comprising the BNB, and highlights the pathology and pathomechanisms of BNB breakdown in immune mediated neuropathies. The human immortalised cell lines of BNB origin established in our laboratory will facilitate the future development of BNB research. Potential therapeutic strategies for immune mediated neuropathies manipulating the BNB are also discussed.

Micropallet arrays for the capture, isolation and culture of circulating tumor cells from whole blood of mice engrafted with primary human pancreatic adenocarcinoma.

PubMed

Gach, Philip C; Attayek, Peter J; Whittlesey, Rebecca L; Yeh, Jen Jen; Allbritton, Nancy L

2014-04-15

Circulating tumor cells (CTCs) are important biomarkers of cancer progression and metastatic potential. The rarity of CTCs in peripheral blood has driven the development of technologies to isolate these tumor cells with high specificity; however, there are limited techniques available for isolating target CTCs following enumeration. A strategy is described to capture and isolate viable tumor cells from whole blood using an array of releasable microstructures termed micropallets. Specific capture of nucleated cells or cells expressing epithelial cell adhesion molecules (EpCAM) was achieved by functionalizing micropallet surfaces with either fibronectin, Matrigel or anti-EpCAM antibody. Surface grafting of poly(acrylic acid) followed by covalent binding of protein A/G enabled efficient capture of EpCAM antibody on the micropallet surface. MCF-7 cells, a human breast adenocarcinoma, were retained on the array surface with 90±8% efficiency when using an anti-EpCAM-coated array. To demonstrate the efficiency of tumor cell retention on micropallet arrays in the presence of blood, MCF-7 cells were mixed into whole blood and added to small arrays (71 mm(2)) coated with fibronectin, Matrigel or anti-EpCAM. These approaches achieved MCF-7 cell capture from ≤10 µL of whole blood with efficiencies greater than 85%. Furthermore, MCF-7 cells intermixed with 1 mL blood and loaded onto large arrays (7171 mm(2)) were captured with high efficiencies (≥97%), could be isolated from the array by a laser-based approach and were demonstrated to yield a high rate of colony formation (≥85%) after removal from the array. Clinical utility of this technology was shown through the capture, isolation and successful culture of CTCs from the blood of mice engrafted with primary human pancreatic tumors. Direct capture and isolation of living tumor cells from blood followed by analysis or culture will be a valuable tool for cancer cell characterization. © 2013 Elsevier B.V. All rights reserved.

Genetics Home Reference: pyruvate kinase deficiency

MedlinePlus

... glucose is broken down to produce adenosine triphosphate (ATP), the cell's main energy source. PKLR gene mutations ... pyruvate kinase enzyme function, causing a shortage of ATP in red blood cells and increased levels of ...

Two ways to feel the pressure: an endothelial Ca(2+) entry channel with dual mechanosensitivity.

PubMed

Groschner, Klaus

2002-01-01

One impressive function of the vascular endothelium is its ability to adjust the release of vasoactive mediators such as NO and PGI(2) almost instantaneously to changes in blood flow or blood pressure. Besides this fast feedback response to hemodynamic alterations, the endothelium is subject to long-term adaptations that are crucial for prevention of pathological processes such as atherogenesis. Among the various signals that are sensed by endothelial cells, mechanical forces which arise from pulsatile blood flow are probably most important for fast as well as long-term control of blood vessel function by the endothelium.

The resident macrophages in murine pancreatic islets are constantly probing their local environment, capturing beta cell granules and blood particles.

PubMed

Zinselmeyer, Bernd H; Vomund, Anthony N; Saunders, Brian T; Johnson, Michael W; Carrero, Javier A; Unanue, Emil R

2018-06-01

We studied here the interactions between the resident macrophages of pancreatic islets with beta cells and the blood vasculature. We also examined the immunological consequences of such interactions. Islets were isolated from C57BL/6 mice expressing CX3C motif chemokine receptor 1-green fluorescent protein (CX3CR-GFP) and examined live by two-photon microscopy. Islets were also examined by electron microscopy to study the relationship of the intra-islet macrophages with the beta cells. In NOD.Rag1 -/- mice and young (non-diabetic) male mice, the acquisition of beta cell granules was tested functionally by probing with CD4 + T cells directed against insulin epitopes. Two-photon microscopy showed that the islet resident macrophages were in close contact with blood vessels and had extensive filopodial activity. Some filopodia had direct access to the vessel lumen and captured microparticles. Addition of glucose at high concentration reduced the degree of filopodia sampling of islets. This finding applied to in vivo injection of glucose or to in vitro cultures. Ultrastructural examination showed the close contacts of macrophages with beta cells. Such macrophages contained intact dense core granules. Functional studies in NOD mice indicated that the macrophages presented insulin peptides to insulin-reactive T cells. Presentation was increased after glucose challenge either ex vivo or after an in vivo pulse. In agreement with the morphological findings, presentation was not affected by insulin receptor blockade. Islet resident macrophages are highly active, sampling large areas of the islets and blood contents and capturing beta cell granules. After such interactions, macrophages present immunogenic insulin to specific autoreactive T cells.

White blood cell counting system

NASA Technical Reports Server (NTRS)

1972-01-01

The design, fabrication, and tests of a prototype white blood cell counting system for use in the Skylab IMSS are presented. The counting system consists of a sample collection subsystem, sample dilution and fluid containment subsystem, and a cell counter. Preliminary test results show the sample collection and the dilution subsystems are functional and fulfill design goals. Results for the fluid containment subsystem show the handling bags cause counting errors due to: (1) adsorption of cells to the walls of the container, and (2) inadequate cleaning of the plastic bag material before fabrication. It was recommended that another bag material be selected.

Measurement of exercise-induced oxidative stress in lymphocytes.

PubMed

Turner, James E; Bosch, Jos A; Aldred, Sarah

2011-10-01

Vigorous exercise is associated with oxidative stress, a state that involves modifications to bodily molecules due to release of pro-oxidant species. Assessment of such modifications provides non-specific measures of oxidative stress in human tissues and blood, including circulating lymphocytes. Lymphocytes are a very heterogeneous group of white blood cells, consisting of subtypes that have different functions in immunity. Importantly, exercise drastically changes the lymphocyte composition in blood by increasing the numbers of some subsets, while leaving other cells unaffected. This fact may imply that observed changes in oxidative stress markers are confounded by changes in lymphocyte composition. For example, lymphocyte subsets may differ in exposure to oxidative stress because of subset differences in cell division and the acquisition of cytotoxic effector functions. The aim of the present review is to raise awareness of interpretational issues related to the assessment of oxidative stress in lymphocytes with exercise and to address the relevance of lymphocyte subset phenotyping in these contexts.

Serotonergic raphe magnus cell discharge reflects ongoing autonomic and respiratory activities.

PubMed

Mason, Peggy; Gao, Keming; Genzen, Jonathan R

2007-10-01

Serotonergic cells are located in a restricted number of brain stem nuclei, send projections to virtually all parts of the CNS, and are critical to normal brain function. They discharge tonically at a rate modulated by the sleep-wake cycle and, in the case of medullary serotonergic cells in raphe magnus and the adjacent reticular formation (RM), are excited by cold challenge. Yet, beyond behavioral state and cold, endogenous factors that influence serotonergic cell discharge remain largely mysterious. The present study in the anesthetized rat investigated predictors of serotonergic RM cell discharge by testing whether cell discharge correlated to three rhythms observed in blood pressure recordings that averaged >30 min in length. A very slow frequency rhythm with a period of minutes, a respiratory rhythm, and a cardiac rhythm were derived from the blood pressure recording. Cross-correlations between each of the derived rhythms and cell activity revealed that the discharge of 38 of the 40 serotonergic cells studied was significantly correlated to the very slow and/or respiratory rhythms. Very few serotonergic cells discharged in relation to the cardiac cycle and those that did, did so weakly. The correlations between serotonergic cell discharge and the slow and respiratory rhythms cannot arise from baroreceptive input. Instead we hypothesize that they are by-products of ongoing adjustments to homeostatic functions that happen to alter blood pressure. Thus serotonergic RM cells integrate information about multiple homeostatic activities and challenges and can consequently modulate spinal processes according to the most pressing need of the organism.

Mesenchymal stem cells attenuate blood-brain barrier leakage after cerebral ischemia in mice.

PubMed

Cheng, Zhuo; Wang, Liping; Qu, Meijie; Liang, Huaibin; Li, Wanlu; Li, Yongfang; Deng, Lidong; Zhang, Zhijun; Yang, Guo-Yuan

2018-05-03

Ischemic stroke induced matrixmetallo-proteinase-9 (MMP-9) upregulation, which increased blood-brain barrier permeability. Studies demonstrated that mesenchymal stem cell therapy protected blood-brain barrier disruption from several cerebrovascular diseases. However, the underlying mechanism was largely unknown. We therefore hypothesized that mesenchymal stem cells reduced blood-brain barrier destruction by inhibiting matrixmetallo-proteinase-9 and it was related to intercellular adhesion molecule-1 (ICAM-1). Adult ICR male mice (n = 118) underwent 90-min middle cerebral artery occlusion and received 2 × 10 5 mesenchymal stem cell transplantation. Neurobehavioral outcome, infarct volume, and blood-brain barrier permeability were measured after ischemia. The relationship between myeloperoxidase (MPO) activity and ICAM-1 release was further determined. We found that intracranial injection of mesenchymal stem cells reduced infarct volume and improved behavioral function in experimental stroke models (p < 0.05). IgG leakage, tight junction protein loss, and inflammatory cytokines IL-1β, IL-6, and TNF-α reduced in mesenchymal stem cell-treated mice compared to the control group following ischemia (p < 0.05). After transplantation, MMP-9 was decreased in protein and activity levels as compared with controls (p < 0.05). Furthermore, myeloperoxidase-positive cells and myeloperoxidase activity were decreased in mesenchymal stem cell-treated mice (p < 0.05). The results showed that mesenchymal stem cell therapy attenuated blood-brain barrier disruption in mice after ischemia. Mesenchymal stem cells attenuated the upward trend of MMP-9 and potentially via downregulating ICAM-1 in endothelial cells. Adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) pathway may influence MMP-9 expression of neutrophils and resident cells, and ICAM-1 acted as a key factor in the paracrine actions of mesenchymal stem cell.

In vitro studies of the blood-brain barrier using isolated brain capillaries and cultured endothelial cells.

PubMed

Goldstein, G W; Betz, A L; Bowman, P D; Dorovini-Zis, K

1986-01-01

The endothelial cells in brain capillaries are the anatomic site of the blood-brain barrier. To learn more about the biology of these specialized cells, we developed methods to prepare suspensions of purified brain microvessels as well as primary cultures of endothelial cells in monolayer. These two preparations allow for direct investigation of the metabolism, transport properties, and receptor content of the brain capillary. We used isolated brain microvessels to study distribution of membrane carriers between the luminal and the abluminal plasma membrane of endothelial cells. We found that Na+K+-ATPase and the A-system amino-acid transport system are located predominantly on the abluminal surface of brain capillary endothelial cells. This distribution of transport carriers is consistent with the low permeability of potassium and small neutral amino acids in the blood-to-brain direction. It suggests, however, that both solutes can be actively transported across brain capillaries from the brain interstitial fluid to the blood. In tissue culture, the endothelial cells form continuous tight junctions with their neighbors. This results in a cellular layer impermeable to protein tracers. When exposed to hyperosmolar solutions, in an attempt to mimic the conditions that open the blood-brain barrier in vivo, we found a reversible separation of the tight junctions between contiguous endothelial cells. No indication of activation of pinocytosis was observed. In vitro systems provide a novel approach for studying the function of the blood-brain barrier and allow for observations not possible with intact animals.

In vivo blockade of the PD-1 pathway using soluble rPD-1-Fc enhances CD4+ and CD8+ T cell responses but has limited clinical benefit

PubMed Central

Amancha, Praveen K.; Hong, Jung Joo; Rogers, Kenneth; Ansari, Aftab A.; Villinger, Francois

2013-01-01

The PD-1/PD-Ligand pathway has been shown to limit cell mediated effector functions during chronic viral infections impeding clearance of pathogens. As a strategy to reverse this exhaustion and increase T cell poly-functionality, PD-1 ligands were blocked in vivo using a recombinant macaque PD1-Fc fusion protein (rPD-1-Fc) in SIVmac239 infected rhesus macaques during the early chronic phase of infection, either alone or in combination with ART. In vitro blockade showed improvement of antigen specific CD4+ and CD8+ T cells from monkeys chronically infected with SIV. Of note, a prolonged 5-day blockade in culture was beneficial for both gag specific CD4+ and CD8+ T cells based on proliferation and dual cytokine production. While the in vivo administration of a recombinant rhesus PD-1 Fc fusion protein (rPD-1-Fc) induced enhanced SIV specific CD4 and CD8 T cell proliferation both in the blood and gut, it failed to alter plasma viremia. However, rPD-1-Fc administration in the context of ART interruption induced a significant delay of viral load rebound. In addition, rPD-1-Fc administration in MamuA*001+ monkeys led to both an increase in the frequencies and Ki67 expression of GagCM9+ CD8+ T cells in the blood and rectal mucosa and poly-functionality of GagCM9+ CD8+ T cells in blood. In conclusion, however, our data suggest that PD-1/PD-Ligand blockade using soluble rPD-1-Fc instead of anti-PD1 Mab, while effective in rescuing the effector function of SIV-specific CD4+ and CD8+ T cells during the early chronic phase of infection, has limited clinical benefit. PMID:24227774

Deficiency of bone marrow beta3-integrin enhances non-functional neovascularization.

PubMed

Watson, Alan R; Pitchford, Simon C; Reynolds, Louise E; Direkze, Natalie; Brittan, Mairi; Alison, Malcolm R; Rankin, Sara; Wright, Nicholas A; Hodivala-Dilke, Kairbaan M

2010-03-01

beta3-Integrin is a cell surface adhesion and signalling molecule important in the regulation of tumour angiogenesis. Mice with a global deficiency in beta3-integrin show increased pathological angiogenesis, most likely due to increased vascular endothelial growth factor receptor 2 expression on beta3-null endothelial cells. Here we transplanted beta3-null bone marrow (BM) into wild-type (WT) mice to dissect the role of BM beta3-integrin deficiency in pathological angiogenesis. Mice transplanted with beta3-null bone marrow show significantly enhanced angiogenesis in subcutaneous B16F0 melanoma and Lewis lung carcinoma (LLC) cell models and in B16F0 melanoma lung metastasis when compared with tumours grown in mice transplanted with WT bone marrow. The effect of bone marrow beta3-integrin deficiency was also assessed in the RIPTAg mouse model of pancreatic tumour growth. Again, angiogenesis in mice lacking BM beta3-integrin was enhanced. However, tumour weight between the groups was not significantly altered, suggesting that the enhanced blood vessel density in the mice transplanted with beta3-null bone marrow was not functional. Indeed, we demonstrate that in mice transplanted with beta3-null bone marrow a significant proportion of tumour blood vessels are non-functional when compared with tumour blood vessels in WT-transplanted controls. Furthermore, beta3-null-transplanted mice showed an increased angiogenic response to VEGF in vivo when compared with WT-transplanted animals. BM beta3-integrin deficiency affects the mobilization of progenitor cells to the peripheral circulation. We show that VEGF-induced mobilization of endothelial progenitor cells is enhanced in mice transplanted with beta3-null bone marrow when compared with WT-transplanted controls, suggesting a possible mechanism underlying the increased blood vessel density seen in beta3-null-transplanted mice. In conclusion, although BM beta3-integrin is not required for pathological angiogenesis, our studies demonstrate a role for BM beta3-integrin in VEGF-induced mobilization of bone marrow-derived cells to the peripheral circulation and for the functionality of those vessels in which BM-derived cells become incorporated.

“You Shall Not Pass”—tight junctions of the blood brain barrier

PubMed Central

Bauer, Hans-Christian; Krizbai, István A.; Bauer, Hannelore; Traweger, Andreas

2014-01-01

The structure and function of the barrier layers restricting the free diffusion of substances between the central nervous system (brain and spinal cord) and the systemic circulation is of great medical interest as various pathological conditions often lead to their impairment. Excessive leakage of blood-borne molecules into the parenchyma and the concomitant fluctuations in the microenvironment following a transient breakdown of the blood-brain barrier (BBB) during ischemic/hypoxic conditions or because of an autoimmune disease are detrimental to the physiological functioning of nervous tissue. On the other hand, the treatment of neurological disorders is often hampered as only minimal amounts of therapeutic agents are able to penetrate a fully functional BBB or blood cerebrospinal fluid barrier. An in-depth understanding of the molecular machinery governing the establishment and maintenance of these barriers is necessary to develop rational strategies allowing a controlled delivery of appropriate drugs to the CNS. At the basis of such tissue barriers are intimate cell-cell contacts (zonulae occludentes, tight junctions) which are present in all polarized epithelia and endothelia. By creating a paracellular diffusion constraint TJs enable the vectorial transport across cell monolayers. More recent findings indicate that functional barriers are already established during development, protecting the fetal brain. As an understanding of the biogenesis of TJs might reveal the underlying mechanisms of barrier formation during ontogenic development numerous in vitro systems have been developed to study the assembly and disassembly of TJs. In addition, monitoring the stage-specific expression of TJ-associated proteins during development has brought much insight into the “developmental tightening” of tissue barriers. Over the last two decades a detailed molecular map of transmembrane and cytoplasmic TJ-proteins has been identified. These proteins not only form a cell-cell adhesion structure, but integrate various signaling pathways, thereby directly or indirectly impacting upon processes such as cell-cell adhesion, cytoskeletal rearrangement, and transcriptional control. This review will provide a brief overview on the establishment of the BBB during embryonic development in mammals and a detailed description of the ultrastructure, biogenesis, and molecular composition of epithelial and endothelial TJs will be given. PMID:25520612

Molecular Parallels between Neural and Vascular Development

PubMed Central

Eichmann, Anne; Thomas, Jean-Léon

2013-01-01

The human central nervous system (CNS) features a network of ∼400 miles of blood vessels that receives >20% of the body’s cardiac output and uses most of its blood glucose. Many human diseases, including stroke, retinopathy, and cancer, are associated with the biology of CNS blood vessels. These vessels originate from extrinsic cell populations, including endothelial cells and pericytes that colonize the CNS and interact with glia and neurons to establish the blood–brain barrier and control cerebrovascular exchanges. Neurovascular interactions also play important roles in adult neurogenic niches, which harbor a unique population of neural stem cells that are intimately associated with blood vessels. We here review the cellular and molecular mechanisms required to establish the CNS vascular network, with a special focus on neurovascular interactions and the functions of vascular endothelial growth factors. PMID:23024177

Airway and alveolar nitric oxide production, lung function, and pulmonary blood flow in sickle cell disease.

PubMed

Lunt, Alan; Ahmed, Na'eem; Rafferty, Gerrard F; Dick, Moira; Rees, David; Height, Sue; Thein, Swee Lay; Greenough, Anne

2016-02-01

Children with sickle cell disease (SCD) often have obstructive lung function abnormalities which could be due to asthma or increased pulmonary blood volume; it is important to determine the underlying mechanism to direct appropriate treatment. In asthmatics, exhaled nitric oxide (FeNO) is elevated. FeNO, however, can also be raised due to increased alveolar production. Our aim, therefore, was to determine if airway or alveolar NO production differed between SCD children and ethnic and age-matched controls. Lung function, airway NO flux and alveolar NO production, and effective pulmonary blood flow were assessed in 18 SCD children and 18 ethnic and age-matched controls. The SCD children compared to the controls had a higher respiratory system resistance (P = 0.0008), alveolar NO production (P = 0.0224), and pulmonary blood flow (P < 0.0001), but not airway NO flux. There was no significant correlation between FeNO and respiratory system resistance in either group, but in the SCD children, there were correlations between alveolar NO production (P = 0.0006) and concentration (P < 0.0001) and pulmonary blood flow. Airway NO flux was not elevated in the SCD children nor correlated with airways obstruction, suggesting that airways obstruction, at least in some SCD children, is not due to asthma.

Endothelial progenitor cells bind and inhibit platelet function and thrombus formation.

PubMed

Abou-Saleh, Haissam; Yacoub, Daniel; Théorêt, Jean-François; Gillis, Marc-Antoine; Neagoe, Paul-Eduard; Labarthe, Benoit; Théroux, Pierre; Sirois, Martin G; Tabrizian, Maryam; Thorin, Eric; Merhi, Yahye

2009-12-01

Interactions of endothelial progenitor cells (EPCs) with vascular and blood cells contribute to vascular homeostasis. Although platelets promote the homing of EPCs to sites of vascular injury and their differentiation into endothelial cells, the functional consequences of such interactions on platelets remain unknown. Herein, we addressed the interactions between EPCs and platelets and their impact on platelet function and thrombus formation. Cultured on fibronectin in conditioned media, human peripheral blood mononuclear cells differentiated, within 10 days of culture, into EPCs, which uptake acetylated low-density lipoprotein, bind ulex-lectin, lack monocyte/leukocyte markers (CD14, P-selectin glycoprotein ligand-1, L-selectin), express progenitor/endothelial markers (CD34, vascular endothelial growth factor receptor-2, von Willebrand factor, and vascular endothelial cadherin), and proliferate in culture. These EPCs bound activated platelets via CD62P and inhibited its translocation, glycoprotein IIb/IIIa activation, aggregation, and adhesion to collagen, mainly via prostacyclin secretion. Indeed, this was associated with upregulation of cyclooxygenase-2 and inducible nitric oxide synthase. However, the effects on platelets in vitro were reversed by cyclooxygenase and cyclooxygenase-2 inhibition but not by nitric oxide or inducible nitric oxide synthase inhibition. Moreover, in a ferric chloride-induced murine arterial thrombosis model, injection of EPCs led to their incorporation into sites of injury and impaired thrombus formation, leading to an incomplete occlusion with 50% residual flow. Peripheral blood mononuclear cell-derived EPCs bind platelets via CD62P and inhibit platelet activation, aggregation, adhesion to collagen, and thrombus formation, predominantly via upregulation of cyclooxygenase-2 and secretion of prostacyclin. These findings add new insights into the biology of EPCs and define their potential roles in regulating platelet function and thrombosis.

Renal intercalated cells and blood pressure regulation

PubMed Central

Wall, Susan M.

2017-01-01

Type B and non-A, non-B intercalated cells are found within the connecting tubule and the cortical collecting duct. Of these cell types, type B intercalated cells are known to mediate Cl− absorption and HCO3− secretion largely through pendrin-dependent Cl−/HCO3− exchange. This exchange is stimulated by angiotensin II administration and is also stimulated in models of metabolic alkalosis, for instance after aldosterone or NaHCO3 administration. In some rodent models, pendrin-mediated HCO3− secretion modulates acid-base balance. However, the role of pendrin in blood pressure regulation is likely of more physiological or clinical significance. Pendrin regulates blood pressure not only by mediating aldosterone-sensitive Cl− absorption, but also by modulating the aldosterone response for epithelial Na+ channel (ENaC)-mediated Na+ absorption. Pendrin regulates ENaC through changes in open channel of probability, channel surface density, and channels subunit total protein abundance. Thus, aldosterone stimulates ENaC activity through both direct and indirect effects, the latter occurring through its stimulation of pendrin expression and function. Therefore, pendrin contributes to the aldosterone pressor response. Pendrin may also modulate blood pressure in part through its action in the adrenal medulla, where it modulates the release of catecholamines, or through an indirect effect on vascular contractile force. This review describes how aldosterone and angiotensin II-induced signaling regulate pendrin and the contributory role of pendrin in distal nephron function and blood pressure. PMID:29285423

NMDAR antagonists for the treatment of diabetes mellitus-Current status and future directions.

PubMed

Welters, Alena; Klüppel, Carina; Mrugala, Jessica; Wörmeyer, Laura; Meissner, Thomas; Mayatepek, Ertan; Heiss, Christian; Eberhard, Daniel; Lammert, Eckhard

2017-09-01

Diabetes mellitus is characterized by chronically elevated blood glucose levels accelerated by a progressive decline of insulin-producing β-cells in the pancreatic islets. Although medications are available to transiently adjust blood glucose to normal levels, the effects of current drugs are limited when it comes to preservation of a critical mass of functional β-cells to sustainably maintain normoglycemia. In this review, we recapitulate recent evidence on the role of pancreatic N-methyl-D-aspartate receptors (NMDARs) in β-cell physiology, and summarize effects of morphinan-based NMDAR antagonists that are beneficial for insulin secretion, glucose tolerance and islet cell survival. We further discuss NMDAR-mediated molecular pathways relevant for neuronal cell survival, which may also be important for the preservation of β-cell function and mass. Finally, we summarize the literature for evidence on the role of NMDARs in the development of diabetic long-term complications, and highlight beneficial pharmacologic aspects of NMDAR antagonists in diabetic nephropathy, retinopathy as well as neuropathy. © 2017 John Wiley & Sons Ltd.

Improved priming for mobilization of and optimal timing for harvest of peripheral blood stem cells.

PubMed

Knudsen, L M; Gaarsdal, E; Jensen, L; Nielsen, K J; Nikolaisen, K; Johnsen, H E

1996-08-01

The time of stem cell harvest and the mobilization regimen may play important roles in terms of achieving adequate numbers of stem cells by leukapheresis. To optimize the timing of leukapheresis, we have determined simultaneously the number of CD34+ cells in the peripheral blood as well as in the leukapheresis product of 214 apheresis procedures performed in 66 unselected patients with malignant hematologic diseases and solid tumors. A significant correlation between the number of CD34+ cells in peripheral blood and the leukapheresis product (R = 0.8) was found. The presence of more than 20 x 10(3)/ml blood CD34+ cells gave a sufficient yield (> or = 1.0 x 10(6) CD34+ cells/kg) in 81% of the cases. In an attempt to compare two priming regimens, we performed leukapheresis twice in 12 patients with stable disease. In the first sequence, stem cells were mobilized with rhG-CSF (10 micrograms/kg/day) alone and, in the second sequence, with cyclophosphamide (4 g/m2) plus rhG-CSF. A significantly higher yield of CD34+ cells and a better correlation between CD34+ cells in the peripheral blood and the leukapheresis product were found after priming with high-dose cyclophosphamide plus rhG-CSF, compared with priming with rhG-CSF alone. In a multivariate analysis, three factors were found to correlate with the yield of CD34+ cells, namely prior chemotherapy, bone marrow function, and the mobilization regimen. The use of cyclophosphamide priming improves CD34+ mobilization, and the introduction of blood CD34+ level optimizes the timing for harvest of stem cells, which should be performed early during treatment of malignancies.

Stable chromosomal aberrations in haemopoietic stem cells in the blood of radiation accident victims.

PubMed

Kreja, L; Greulich, K M; Fliedner, T M; Heinze, B

1999-10-01

The detection of long-term persistent chromosome aberrations in circulating haemopoietic stem cells after accidental radiation exposure. Peripheral blood samples from highly exposed persons were collected 7-25 years after the radiation accidents in Moscow (1971), Kazan (1975) and Chernobyl (1996). Haemopoietic blood stem cells were analysed when investigating individual colonies derived from haemopoietic progenitor cells: burst-forming units-erythroid (BFU-E), granulocyte-macrophage-colony-forming cells (GM-CFC) and multipotent granulocyte-erythrocyte-macrophage- megakaryocyte-colony-forming cells (GEMM-CFC). Colony formation was obtained in methylcellulose cultures. Chromosome preparations in single colonies were performed using a microtechnique. Nine patients were investigated at 1 to 4 follow-up time points after radiation exposure. Three hundred and thirty-four single colonies were analyzed resulting in 1375 mitoses. It was found that colonies showed chromosome aberrations (ChA) up to 25 years after radiation exposure by classical cytogenetics and by fluorescence in situ hybridization (FISH). Stable aberrations were detected in 21% of colonies. They were clonal in 19% of colonies, i.e. the same abnormality was found in all cells derived from a single colony. In 2% of colonies ChA were stable but non-clonal; unstable ChA were not observed. The results indicate that blood-derived haemopoietic stem cells may serve as a biological indicator to detect radiation-induced ChA. Since they are considered to be in dynamic and functional exchange with stem cells in the medullary sites of blood cell formation such as bone marrow, the use of blood stem cells as a marker of radiation effects should be explored to assess the repair status of the stem cell pool as such.

Covalent Binding of Heparin to Functionalized PET Materials for Improved Haemocompatibility

PubMed Central

Kolar, Metod; Mozetič, Miran; Stana-Kleinschek, Karin; Fröhlich, Mirjam; Turk, Boris; Vesel, Alenka

2015-01-01

The hemocompatibility of vascular grafts made from poly(ethylene terephthalate) (PET) is insufficient due to the rapid adhesion and activation of blood platelets that occur upon incubation with whole blood. PET polymer was treated with NHx radicals created by passing ammonia through gaseous plasma formed by a microwave discharge, which allowed for functionalization with amino groups. X-ray photoelectron spectroscopy characterization using derivatization with 4-chlorobenzaldehyde indicated that approximately 4% of the –NH2 groups were associated with the PET surface after treatment with the gaseous radicals. The functionalized polymers were coated with an ultra-thin layer of heparin and incubated with fresh blood. The free-hemoglobin technique, which is based on the haemolysis of erythrocytes, indicated improved hemocompatibility, which was confirmed by imaging the samples using confocal optical microscopy. A significant decrease in number of adhered platelets was observed on such samples. Proliferation of both human umbilical vein endothelial cells and human microvascular endothelial cells was enhanced on treated polymers, especially after a few hours of cell seeding. Thus, the technique represents a promising substitute for wet-chemical modification of PET materials prior to coating with heparin. PMID:28788016

[Significance of considering hemoglobin derivatives and acid-base balance in the evaluation of the blood oxygen-transport system].

PubMed

Matiushichev, V B; Shamratova, V G; Krapivko, Iu K

2009-12-01

Factor analysis was used to study the pattern of relationships of a number of hematological parameters in clinically healthy young subjects and in patients with moderate anemia. The level of total hemoglobin and the concentration of red blood cells were ascertained to control blood oxygen-transporting function in not full measure and these might be referred to as basic characteristics only conventionally. To clarify the picture, these criteria should be supplemented by the information on other parameters. It is concluded that the introduction of the ratio of a number of hemoglobin derivatives, blood oxygen regimen and acid-base balance can substantially increase the validity of clinical opinions as to this blood function.

Acquired Downregulation of Donor-Specific Antibody Production After ABO-Incompatible Kidney Transplantation.

PubMed

Tasaki, M; Saito, K; Nakagawa, Y; Imai, N; Ito, Y; Aoki, T; Kamimura, M; Narita, I; Tomita, Y; Takahashi, K

2017-01-01

The mechanism of long-term B cell immunity against donor blood group antigens in recipients who undergo ABO-incompatible (ABOi) living-donor kidney transplantation (LKTx) is unknown. To address this question, we evaluated serial anti-A and anti-B antibody titers in 50 adult recipients. Donor-specific antibody titers remained low (≤1:4) in 42 recipients (84%). However, antibodies against nondonor blood group antigens were continuously produced in recipients with blood type O. We stimulated recipients' peripheral blood mononuclear cells in vitro to investigate whether B cells produced antibodies against donor blood group antigens in the absence of graft adsorption in vivo. Antibodies in cell culture supernatant were measured using specific enzyme-linked immunosorbent assays (ELISAs). Thirty-five healthy volunteers and 57 recipients who underwent ABO-compatible LKTx served as controls. Antibody production in vitro against donor blood group antigens by cells from ABOi LKTx patients was lower than in the control groups. Immunoglobulin deposits were undetectable in biopsies of grafts of eight recipients with low antibody titers (≤1:4) after ABOi LKTx. One patient with blood type A1 who received a second ABOi LKTx from a type B donor did not produce B-specific antibodies. These findings suggest diminished donor-specific antibody production function in the setting of adult ABOi LKTx. © Copyright 2016 The American Society of Transplantation and the American Society of Transplant Surgeons.

iss056e009809

NASA Image and Video Library

2018-06-11

iss056e009809 (June 11, 2018) --- Expedition 56 Flight Engineer Serena Auñón-Chancellor of NASA is pictured in the Destiny laboratory module with gear from the Marrow investigation. She was collecting breath samples to analyze and measure red blood cell function to help doctors understand how blood cell production is altered in microgravity. Results may improve the health of astronauts on long-term missions and help patients on Earth with mobility and aging issues.

Acute lead poisoning in two users of illicit methamphetamine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Allcott, J.V. III; Barnhart, R.A.; Mooney, L.A.

1987-07-31

Acute lead poisoning can present a difficult diagnostic dilemma, with symptoms that mimic those of hepatitis, nephritis, and encephalopathy. The authors report two cases in intravenous methamphetamine users who presented with abnormal liver function values, low hematocrit values, basophilic stippling of red blood cells, and elevated blood lead levels. Both patients excreted large amounts of lead in their urine after treatment with edetic acid, followed by resolution of their symptoms. Lead contamination was proved in one drug sample. Basophilic stippling of the red blood cells was the one key laboratory result that led to the definitive diagnosis in both cases.

The meningeal lymphatic system: a route for HIV brain migration?

PubMed

Lamers, Susanna L; Rose, Rebecca; Ndhlovu, Lishomwa C; Nolan, David J; Salemi, Marco; Maidji, Ekaterina; Stoddart, Cheryl A; McGrath, Michael S

2016-06-01

Two innovative studies recently identified functional lymphatic structures in the meninges that may influence the development of HIV-associated neurological disorders (HAND). Until now, blood vessels were assumed to be the sole transport system by which HIV-infected monocytes entered the brain by bypassing a potentially hostile blood-brain barrier through inflammatory-mediated semi-permeability. A cascade of specific chemokine signals promote monocyte migration from blood vessels to surrounding brain tissues via a well-supported endothelium, where the cells differentiate into tissue macrophages capable of productive HIV infection. Lymphatic vessels on the other hand are more loosely organized than blood vessels. They absorb interstitial fluid from bodily tissues where HIV may persist and exchange a variety of immune cells (CD4(+) T cells, monocytes, macrophages, and dendritic cells) with surrounding tissues through discontinuous endothelial junctions. We propose that the newly discovered meningeal lymphatics are key to HIV migration among viral reservoirs and brain tissue during periods of undetectable plasma viral loads due to suppressive combinational antiretroviral therapy, thus redefining the migration process in terms of a blood-lymphatic transport system.

Circulating and Tissue-Resident CD4+ T Cells With Reactivity to Intestinal Microbiota Are Abundant in Healthy Individuals and Function Is Altered During Inflammation.

PubMed

Hegazy, Ahmed N; West, Nathaniel R; Stubbington, Michael J T; Wendt, Emily; Suijker, Kim I M; Datsi, Angeliki; This, Sebastien; Danne, Camille; Campion, Suzanne; Duncan, Sylvia H; Owens, Benjamin M J; Uhlig, Holm H; McMichael, Andrew; Bergthaler, Andreas; Teichmann, Sarah A; Keshav, Satish; Powrie, Fiona

2017-11-01

Interactions between commensal microbes and the immune system are tightly regulated and maintain intestinal homeostasis, but little is known about these interactions in humans. We investigated responses of human CD4 + T cells to the intestinal microbiota. We measured the abundance of T cells in circulation and intestinal tissues that respond to intestinal microbes and determined their clonal diversity. We also assessed their functional phenotypes and effects on intestinal resident cell populations, and studied alterations in microbe-reactive T cells in patients with chronic intestinal inflammation. We collected samples of peripheral blood mononuclear cells and intestinal tissues from healthy individuals (controls, n = 13-30) and patients with inflammatory bowel diseases (n = 119; 59 with ulcerative colitis and 60 with Crohn's disease). We used 2 independent assays (CD154 detection and carboxy-fluorescein succinimidyl ester dilution assays) and 9 intestinal bacterial species (Escherichia coli, Lactobacillus acidophilus, Bifidobacterium animalis subsp lactis, Faecalibacterium prausnitzii, Bacteroides vulgatus, Roseburia intestinalis, Ruminococcus obeum, Salmonella typhimurium, and Clostridium difficile) to quantify, expand, and characterize microbe-reactive CD4 + T cells. We sequenced T-cell receptor Vβ genes in expanded microbe-reactive T-cell lines to determine their clonal diversity. We examined the effects of microbe-reactive CD4 + T cells on intestinal stromal and epithelial cell lines. Cytokines, chemokines, and gene expression patterns were measured by flow cytometry and quantitative polymerase chain reaction. Circulating and gut-resident CD4 + T cells from controls responded to bacteria at frequencies of 40-4000 per million for each bacterial species tested. Microbiota-reactive CD4 + T cells were mainly of a memory phenotype, present in peripheral blood mononuclear cells and intestinal tissue, and had a diverse T-cell receptor Vβ repertoire. These cells were functionally heterogeneous, produced barrier-protective cytokines, and stimulated intestinal stromal and epithelial cells via interleukin 17A, interferon gamma, and tumor necrosis factor. In patients with inflammatory bowel diseases, microbiota-reactive CD4 + T cells were reduced in the blood compared with intestine; T-cell responses that we detected had an increased frequency of interleukin 17A production compared with responses of T cells from blood or intestinal tissues of controls. In an analysis of peripheral blood mononuclear cells and intestinal tissues from patients with inflammatory bowel diseases vs controls, we found that reactivity to intestinal bacteria is a normal property of the human CD4 + T-cell repertoire, and does not necessarily indicate disrupted interactions between immune cells and the commensal microbiota. T-cell responses to commensals might support intestinal homeostasis, by producing barrier-protective cytokines and providing a large pool of T cells that react to pathogens. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

Variations of B cell subpopulations in peripheral blood of healthy Mexican population according to age: Relevance for diagnosis of primary immunodeficiencies.

PubMed

Berrón-Ruíz, L; López-Herrera, G; Ávalos-Martínez, C E; Valenzuela-Ponce, C; Ramírez-SanJuan, E; Santoyo-Sánchez, G; Mújica Guzmán, F; Espinosa-Rosales, F J; Santos-Argumedo, L

Peripheral blood B cells include lymphocytes at various stages of differentiation, each with a specific function in the immune response. All these stages show variations in percentage and absolute number throughout human life. The numbers and proportions of B subpopulation are influenced by factors such as gender, age, ethnicity, and lifestyle. This study establishes reference values according to age of peripheral blood B cell subtypes in healthy Mexican population. Peripheral blood from healthy new-borns and adults were analysed for total B cell subpopulations, using surface markers such as CD19, IgM, IgD, CD21, CD24, CD27, and CD38, to identify naïve, memory with and without isotype switch, double-negative, transitional, and plasmablast cells. We observed a significant variation in terms of frequency and absolute counts between all groups analysed. Values from each B cell subpopulation show variations according to age. In order to attempt to elucidate reference values for B cell subpopulation, the present study evaluated a population sample of healthy blood donors from this region. Values reported here can also be used as a tool for diagnosis of diseases in which B cell maturation is affected. Copyright © 2016 SEICAP. Published by Elsevier España, S.L.U. All rights reserved.

Acute Respiratory Distress Syndrome Neutrophils Have a Distinct Phenotype and Are Resistant to Phosphoinositide 3-Kinase Inhibition

PubMed Central

Juss, Jatinder K.; House, David; Amour, Augustin; Begg, Malcolm; Herre, Jurgen; Storisteanu, Daniel M. L.; Hoenderdos, Kim; Bradley, Glyn; Lennon, Mark; Summers, Charlotte; Hessel, Edith M.; Condliffe, Alison

2016-01-01

Rationale: Acute respiratory distress syndrome is refractory to pharmacological intervention. Inappropriate activation of alveolar neutrophils is believed to underpin this disease’s complex pathophysiology, yet these cells have been little studied. Objectives: To examine the functional and transcriptional profiles of patient blood and alveolar neutrophils compared with healthy volunteer cells, and to define their sensitivity to phosphoinositide 3-kinase inhibition. Methods: Twenty-three ventilated patients underwent bronchoalveolar lavage. Alveolar and blood neutrophil apoptosis, phagocytosis, and adhesion molecules were quantified by flow cytometry, and oxidase responses were quantified by chemiluminescence. Cytokine and transcriptional profiling were used in multiplex and GeneChip arrays. Measurements and Main Results: Patient blood and alveolar neutrophils were distinct from healthy circulating cells, with increased CD11b and reduced CD62L expression, delayed constitutive apoptosis, and primed oxidase responses. Incubating control cells with disease bronchoalveolar lavage recapitulated the aberrant functional phenotype, and this could be reversed by phosphoinositide 3-kinase inhibitors. In contrast, the prosurvival phenotype of patient cells was resistant to phosphoinositide 3-kinase inhibition. RNA transcriptomic analysis revealed modified immune, cytoskeletal, and cell death pathways in patient cells, aligning closely to sepsis and burns datasets but not to phosphoinositide 3-kinase signatures. Conclusions: Acute respiratory distress syndrome blood and alveolar neutrophils display a distinct primed prosurvival profile and transcriptional signature. The enhanced respiratory burst was phosphoinositide 3-kinase–dependent but delayed apoptosis and the altered transcriptional profile were not. These unexpected findings cast doubt over the utility of phosphoinositide 3-kinase inhibition in acute respiratory distress syndrome and highlight the importance of evaluating novel therapeutic strategies in patient-derived cells. PMID:27064380

DNA methylation dynamics during in vivo differentiation of blood and skin stem cells

PubMed Central

Bock, Christoph; Beerman, Isabel; Lien, Wen-Hui; Smith, Zachary D.; Gu, Hongcang; Boyle, Patrick; Gnirke, Andreas; Fuchs, Elaine; Rossi, Derrick J.; Meissner, Alexander

2012-01-01

DNA methylation is a mechanism of epigenetic regulation that is common to all vertebrates. Functional studies underscore its relevance for tissue homeostasis, but the global dynamics of DNA methylation during in vivo differentiation remain underexplored. Here we report high-resolution DNA methylation maps of adult stem cell differentiation in mouse, focusing on 19 purified cell populations of the blood and skin lineages. DNA methylation changes were locus-specific and relatively modest in magnitude. They frequently overlapped with lineage-associated transcription factors and their binding sites, suggesting that DNA methylation may protect cells from aberrant transcription factor activation. DNA methylation and gene expression provided complementary information, and combining the two enabled us to infer the cellular differentiation hierarchy of the blood lineage directly from genomic data. In summary, these results demonstrate that in vivo differentiation of adult stem cells is associated with small but informative changes in the genomic distribution of DNA methylation. PMID:22841485

Separation of human CD4+CD39+ T cells by magnetic beads reveals two phenotypically and functionally different subsets

PubMed Central

Schuler, Patrick J.; Harasymczuk, Malgorzata; Schilling, Bastian; Lang, Stephan; Whiteside, Theresa L.

2011-01-01

Objective The ectonucleotidase CD39 is an enzyme involved in adenosine production. Its surface expression on human regulatory T cells (Treg) allows for their flow-cytometry-based isolation from peripheral blood. To further develop and improve this method on a scale supporting translational studies, we introduced capture of CD39+ Treg on magnetic immunobeads. Methods Peripheral blood mononuclear cells (PBMC) obtained from healthy donors were used for negative selection of CD4+ T cells on AutoMACS using antibodies (Abs) specific for all lineage+ cells. CD4+CD39+ Treg were captured by biotin-conjugated anti-CD39 Abs and anti-biotin Ab-coated magnetic beads. Isolated CD4+CD39+ T cells were phenotyped by flow cytometry for Treg-associated markers: CD39, CD73, FOXP3, CD25, CTLA-4, CCR4, CD45RO and CD121a or for the absence of CD127 and CD49d. CFSE-based proliferation assays and ATP hydrolysis were used to measure Treg functions. Results The purity, recovery and viability of the separated CD4+CD39+ T cells were satisfactory. The isolated CD4+CD39+ T cell population consisted of FOXP3+CD25+ T cells which hydrolyzed exogenous ATP and suppressed autologous CD4+ T cell proliferation and of FOXP3negCD25neg T cells without suppressor function. The same two subsets were detectable by flow cytometry in normal PBMC, gating on CD4+CD39+, CD4+CD127neg, CD4+CD49dneg or CD4+CD25high Treg. Conclusion CD4+CD39+ Treg capture on immunobeads led to a discovery of two CD39+ subsets. Similar to CD39+ Treg in the peripheral blood, half of these cells are CD25+FOXP3+ active suppressor cells, while the other half are CD25negFOXP3neg and do not mediate suppression. PMID:21513715

NO supplementation for transfusion medicine and cardiovascular applications.

PubMed

Cabrales, Pedro; Ortiz, Daniel; Friedman, Joel M

Blood transfusions are used to treat reduced O 2 -carrying capacity consequent to anemia. In many cases anemia is caused by a major blood loss, which also creates a state of hypovolemia. Whereas O 2 transport capacity is restored by increasing levels of circulating Hb, transfusion does not resolve the hypoperfusion, the hypoxia and the inflammatory cascades initiated during the anemia and hypovolemia. This explains why blood transfusion is not always an effective treatment and why transfusion of stored blood has been associated with increased morbidity and mortality, especially in patient populations receiving multiple transfusions. Epidemiologic data indicate that adverse events after transfusion are relatively common, having a great impact on the patients outcome and on the costs of public health. In this chapter, we explain why classical transfusion strategies target the reversal of hypoxia only, but do not address the inflammatory cascades initiated during anemic states and the importance of the flow and vascular endothelium interactions. We also establish the relation between red blood cells storage lesions, limited NO bioavailability and transfusion-associated adverse events. Lastly, we explain the potential use of long-lived sources of bioactive NO to reverse the hypoxic inflammatory cascades, promote a sustained increase in tissue perfusion and thereby allow transfusions to achieve their intended goal. The underlying premise is that adverse effects associated with transfusions are intimately linked to vascular dysfunction. Understanding of these mechanisms would lead to novel transfusion medicine strategies to preserve red cell function and to correct for functional changes induced by hemoglobinopathies that affect cell structure and function.

The impact of microglial activation on blood-brain barrier in brain diseases

PubMed Central

da Fonseca, Anna Carolina Carvalho; Matias, Diana; Garcia, Celina; Amaral, Rackele; Geraldo, Luiz Henrique; Freitas, Catarina; Lima, Flavia Regina Souza

2014-01-01

The blood-brain barrier (BBB), constituted by an extensive network of endothelial cells (ECs) together with neurons and glial cells, including microglia, forms the neurovascular unit (NVU). The crosstalk between these cells guarantees a proper environment for brain function. In this context, changes in the endothelium-microglia interactions are associated with a variety of inflammation-related diseases in brain, where BBB permeability is compromised. Increasing evidences indicate that activated microglia modulate expression of tight junctions, which are essential for BBB integrity and function. On the other hand, the endothelium can regulate the state of microglial activation. Here, we review recent advances that provide insights into interactions between the microglia and the vascular system in brain diseases such as infectious/inflammatory diseases, epilepsy, ischemic stroke and neurodegenerative disorders. PMID:25404894

Loss of c-Kit function impairs arteriogenesis in a mouse model of hindlimb ischemia.

PubMed

Hernandez, Diana R; Artiles, Adriana; Duque, Juan C; Martinez, Laisel; Pinto, Mariana T; Webster, Keith A; Velazquez, Omaida C; Vazquez-Padron, Roberto I; Lassance-Soares, Roberta M

2018-04-01

Arteriogenesis is a process whereby collateral vessels remodel usually in response to increased blood flow and/or wall stress. Remodeling of collaterals can function as a natural bypass to alleviate ischemia during arterial occlusion. Here we used a genetic approach to investigate possible roles of tyrosine receptor c-Kit in arteriogenesis. Mutant mice with loss of c-Kit function (Kit W/W-v ), and controls were subjected to hindlimb ischemia. Blood flow recovery was evaluated pre-, post-, and weekly after ischemia. Foot ischemic damage and function were assessed between days 1 to 14 post-ischemia while collaterals remodeling were measured 28 days post-ischemia. Both groups of mice also were subjected to wild type bone marrow cells transplantation 3 weeks before hindlimb ischemia to evaluate possible contributions of defective bone marrow c-Kit expression on vascular recovery. Kit W/W-v mice displayed impaired blood flow recovery, greater ischemic damage and foot dysfunction after ischemia compared to controls. Kit W/W-v mice also demonstrated impaired collateral remodeling consistent with flow recovery findings. Because arteriogenesis is a biological process that involves bone marrow-derived cells, we investigated which source of c-Kit signaling (bone marrow or vascular) plays a major role in arteriogenesis. Kit W/W-v mice transplanted with bone marrow wild type cells exhibited similar phenotype of impaired blood flow recovery, greater tissue ischemic damage and foot dysfunction as nontransplanted Kit W/W-v mice. This study provides evidence that c-Kit signaling is required during arteriogenesis. Also, it strongly suggests a vascular role for c-Kit signaling because rescue of systemic c-Kit activity by bone marrow transplantation did not augment the functional recovery of Kit W/W-v mouse hindlimbs. Copyright © 2017 Elsevier Inc. All rights reserved.

Quantifying effects of cyclic stretch on cell-collagen substrate adhesiveness of vascular endothelial cells.

PubMed

Omidvar, Ramin; Tafazzoli-Shadpour, Mohammad; Mahmoodi-Nobar, Farbod; Azadi, Shohreh; Khani, Mohammad-Mehdi

2018-05-01

Vascular endothelium is continuously subjected to mechanical stimulation in the form of shear forces due to blood flow as well as tensile forces as a consequence of blood pressure. Such stimuli influence endothelial behavior and regulate cell-tissue interaction for an optimized functionality. This study aimed to quantify influence of cyclic stretch on the adhesive property and stiffness of endothelial cells. The 10% cyclic stretch with frequency of 1 Hz was applied to a layer of endothelial cells cultured on a polydimethylsiloxane substrate. Cell-substrate adhesion of endothelial cells was examined by the novel approach of atomic force microscope-based single-cell force spectroscopy and cell stiffness was measured by atomic force microscopy. Furthermore, the adhesive molecular bonds were evaluated using modified Hertz contact theory. Our results show that overall adhesion of endothelial cells with substrate decreased after cyclic stretch while they became stiffer. Based on the experimental results and theoretical modeling, the decrease in the number of molecular bonds after cyclic stretch was quantified. In conclusion, in vitro cyclic stretch caused alterations in both adhesive capacity and elastic modulus of endothelial cells through mechanotransductive pathways as two major determinants of the function of these cells within the cardiovascular system.

Somatic stem cell heterogeneity: diversity in the blood, skin and intestinal stem cell compartments

PubMed Central

Goodell, Margaret A.; Nguyen, Hoang; Shroyer, Noah

2017-01-01

Somatic stem cells replenish many tissues throughout life to repair damage and to maintain tissue homeostasis. Stem cell function is frequently described as following a hierarchical model in which a single master cell undergoes self-renewal and differentiation into multiple cell types and is responsible for most regenerative activity. However, recent data from studies on blood, skin and intestinal epithelium all point to the concomitant action of multiple types of stem cells with distinct everyday roles. Under stress conditions such as acute injury, the surprising developmental flexibility of these stem cells enables them to adapt to diverse roles and to acquire different regeneration capabilities. This paradigm shift raises many new questions about the developmental origins, inter-relationships and molecular regulation of these multiple stem cell types. PMID:25907613

Cellular biophysical markers of hydroxyurea treatment in sickle cell disease

NASA Astrophysics Data System (ADS)

So, Peter T. C.; Hosseini, Poorya; Abidi, Sabia Z.; Du, E.; Papageorgiou, Dimitrios P.; Park, YongKeun; Higgins, John; Kato, Gregory J.; Suresh, Subra; Dao, Ming; Yaqoob, Zahid

2017-04-01

Using a common-path interferometric technique, we measure biomechanical and morphological properties of individual red blood cells in SCD patients as a function of cell density, and investigate the correlation of these biophysical properties with drug intake as well as other clinically measured parameters.

Comparison of the Functional microRNA Expression in Immune Cell Subsets of Neonates and Adults

PubMed Central

Yu, Hong-Ren; Hsu, Te-Yao; Huang, Hsin-Chun; Kuo, Ho-Chang; Li, Sung-Chou; Yang, Kuender D.; Hsieh, Kai-Sheng

2016-01-01

Diversity of biological molecules in newborn and adult immune cells contributes to differences in cell function and atopic properties. Micro RNAs (miRNAs) are reported to involve in the regulation of immune system. Therefore, determining the miRNA expression profile of leukocyte subpopulations is important for understanding immune system regulation. In order to explore the unique miRNA profiling that contribute to altered immune in neonates, we comprehensively analyzed the functional miRNA signatures of eight leukocyte subsets (polymorphonuclear cells, monocytes, CD4+ T cells, CD8+ T cells, natural killer cells, B cells, plasmacytoid dendritic cells, and myeloid dendritic cells) from both neonatal and adult umbilical cord and peripheral blood samples, respectively. We observed distinct miRNA profiles between adult and neonatal blood leukocyte subsets, including unique miRNA signatures for each cell lineage. Leukocyte miRNA signatures were altered after stimulation. Adult peripheral leukocytes had higher let-7b-5p expression levels compared to neonatal cord leukocytes across multiple subsets, irrespective of stimulation. Transfecting neonatal monocytes with a let-7b-5p mimic resulted in a reduction of LPS-induced interleukin (IL)-6 and TNF-α production, while transfection of a let-7b-5p inhibitor into adult monocytes enhanced IL-6 and TNF-α production. With this functional approach, we provide intact differential miRNA expression profiling of specific immune cell subsets between neonates and adults. These studies serve as a basis to further understand the altered immune response observed in neonates and advance the development of therapeutic strategies. PMID:28066425

Genes that mediate breast cancer metastasis to the brain.

PubMed

Bos, Paula D; Zhang, Xiang H-F; Nadal, Cristina; Shu, Weiping; Gomis, Roger R; Nguyen, Don X; Minn, Andy J; van de Vijver, Marc J; Gerald, William L; Foekens, John A; Massagué, Joan

2009-06-18

The molecular basis for breast cancer metastasis to the brain is largely unknown. Brain relapse typically occurs years after the removal of a breast tumour, suggesting that disseminated cancer cells must acquire specialized functions to take over this organ. Here we show that breast cancer metastasis to the brain involves mediators of extravasation through non-fenestrated capillaries, complemented by specific enhancers of blood-brain barrier crossing and brain colonization. We isolated cells that preferentially infiltrate the brain from patients with advanced disease. Gene expression analysis of these cells and of clinical samples, coupled with functional analysis, identified the cyclooxygenase COX2 (also known as PTGS2), the epidermal growth factor receptor (EGFR) ligand HBEGF, and the alpha2,6-sialyltransferase ST6GALNAC5 as mediators of cancer cell passage through the blood-brain barrier. EGFR ligands and COX2 were previously linked to breast cancer infiltration of the lungs, but not the bones or liver, suggesting a sharing of these mediators in cerebral and pulmonary metastases. In contrast, ST6GALNAC5 specifically mediates brain metastasis. Normally restricted to the brain, the expression of ST6GALNAC5 in breast cancer cells enhances their adhesion to brain endothelial cells and their passage through the blood-brain barrier. This co-option of a brain sialyltransferase highlights the role of cell-surface glycosylation in organ-specific metastatic interactions.

Human umbilical cord blood-derived f-macrophages retain pluripotentiality after thrombopoietin expansion

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao Yong; Mazzone, Theodore

2005-11-01

We have previously characterized a new type of stem cell from human peripheral blood, termed fibroblast-like macrophage (f-M{phi}). Here, using umbilical cord blood as a source, we identified cells with similar characteristics including expression of surface markers (CD14, CD34, CD45, CD117, and CD163), phagocytosis, and proliferative capacity. Further, thrombopoietin (TPO) significantly stimulated the proliferation of cord blood-derived f-M{phi} (CB f-M{phi}) at low dosage without inducing a megakaryocytic phenotype. Additional experiments demonstrated that TPO-expanded cord blood-derived f-M{phi} (TCB f-M{phi}) retained their surface markers and differentiation ability. Treatment with vascular endothelial cell growth factor (VEGF) gave rise to endothelial-like cells, expressing Flt-1,more » Flk-1, von Willebrand Factor (vWF), CD31, acetylated low density lipoprotein internalization, and the ability to form endothelial-like cell chains. In the presence of lipopolyssacharide (LPS) and 25 mM glucose, the TCB f-M{phi} differentiated to express insulin mRNA, C-peptide, and insulin. In vitro functional analysis demonstrated that these insulin-positive cells could release insulin in response to glucose and other secretagogues. These findings demonstrate a potential use of CB f-M{phi} and may lead to develop new therapeutic strategy for treating dominant disease.« less

Regulatory T cells ameliorate tissue plasminogen activator-induced brain haemorrhage after stroke.

PubMed

Mao, Leilei; Li, Peiying; Zhu, Wen; Cai, Wei; Liu, Zongjian; Wang, Yanling; Luo, Wenli; Stetler, Ruth A; Leak, Rehana K; Yu, Weifeng; Gao, Yanqin; Chen, Jun; Chen, Gang; Hu, Xiaoming

2017-07-01

Delayed thrombolytic treatment with recombinant tissue plasminogen activator (tPA) may exacerbate blood-brain barrier breakdown after ischaemic stroke and lead to lethal haemorrhagic transformation. The immune system is a dynamic modulator of stroke response, and excessive immune cell accumulation in the cerebral vasculature is associated with compromised integrity of the blood-brain barrier. We previously reported that regulatory T cells, which function to suppress excessive immune responses, ameliorated blood-brain barrier damage after cerebral ischaemia. This study assessed the impact of regulatory T cells in the context of tPA-induced brain haemorrhage and investigated the underlying mechanisms of action. The number of circulating regulatory T cells in stroke patients was dramatically reduced soon after stroke onset (84 acute ischaemic stroke patients with or without intravenous tPA treatment, compared to 115 age and gender-matched healthy controls). Although stroke patients without tPA treatment gradually repopulated the numbers of circulating regulatory T cells within the first 7 days after stroke, post-ischaemic tPA treatment led to sustained suppression of regulatory T cells in the blood. We then used the murine suture and embolic middle cerebral artery occlusion models of stroke to investigate the therapeutic potential of adoptive regulatory T cell transfer against tPA-induced haemorrhagic transformation. Delayed administration of tPA (10 mg/kg) resulted in haemorrhagic transformation in the ischaemic territory 1 day after ischaemia. When regulatory T cells (2 × 106/mouse) were intravenously administered immediately after delayed tPA treatment in ischaemic mice, haemorrhagic transformation was significantly decreased, and this was associated with improved sensorimotor functions. Blood-brain barrier disruption and tight junction damages were observed in the presence of delayed tPA after stroke, but were mitigated by regulatory T cell transfer. Mechanistic studies demonstrated that regulatory T cells completely abolished the tPA-induced elevation of MMP9 and CCL2 after stroke. Using MMP9 and CCL2 knockout mice, we discovered that both molecules partially contributed to the protective actions of regulatory T cells. In an in vitro endothelial cell-based model of the blood-brain barrier, we confirmed that regulatory T cells inhibited tPA-induced endothelial expression of CCL2 and preserved blood-brain barrier integrity after an ischaemic challenge. Lentivirus-mediated CCL2 knockdown in endothelial cells completely abolished the blood-brain barrier protective effect of regulatory T cells in vitro. Altogether, our studies suggest that regulatory T cell adoptive transfer may alleviate thrombolytic treatment-induced haemorrhage in stroke victims. Furthermore, regulatory T cell-afforded protection in the tPA-treated stroke model is mediated by two inhibitory mechanisms involving CCL2 and MMP9. Thus, regulatory T cell adoptive transfer may be useful as a cell-based therapy to improve the efficacy and safety of thrombolytic treatment for ischaemic stroke. © The Author (2017). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Endothelial Notch signalling limits angiogenesis via control of artery formation

PubMed Central

Hasan, Sana S.; Tsaryk, Roman; Lange, Martin; Wisniewski, Laura; Moore, John C.; Lawson, Nathan D.; Wojciechowska, Karolina; Schnittler, Hans; Siekmann, Arndt F.

2017-01-01

Angiogenic sprouting needs to be tightly controlled. It has been suggested that the Notch ligand dll4 expressed in leading tip cells restricts angiogenesis by activating Notch signalling in trailing stalk cells. Here, we show using live imaging in zebrafish that activation of Notch signalling is rather required in tip cells. Notch activation initially triggers expression of the chemokine receptor cxcr4a. This allows for proper tip cell migration and connection to the pre-existing arterial circulation, ultimately establishing functional arterial-venous blood flow patterns. Subsequently, Notch signalling reduces cxcr4a expression, thereby preventing excessive blood vessel growth. Finally, we find that Notch signalling is dispensable for limiting blood vessel growth during venous plexus formation that does not generate arteries. Together, these findings link the role of Notch signalling in limiting angiogenesis to its role during artery formation and provide a framework for our understanding of the mechanisms underlying blood vessel network expansion and maturation. PMID:28714969

Pancreatic islet blood flow and its measurement

PubMed Central

Jansson, Leif; Barbu, Andreea; Bodin, Birgitta; Drott, Carl Johan; Espes, Daniel; Gao, Xiang; Grapensparr, Liza; Källskog, Örjan; Lau, Joey; Liljebäck, Hanna; Palm, Fredrik; Quach, My; Sandberg, Monica; Strömberg, Victoria; Ullsten, Sara; Carlsson, Per-Ola

2016-01-01

Pancreatic islets are richly vascularized, and islet blood vessels are uniquely adapted to maintain and support the internal milieu of the islets favoring normal endocrine function. Islet blood flow is normally very high compared with that to the exocrine pancreas and is autonomously regulated through complex interactions between the nervous system, metabolites from insulin secreting β-cells, endothelium-derived mediators, and hormones. The islet blood flow is normally coupled to the needs for insulin release and is usually disturbed during glucose intolerance and overt diabetes. The present review provides a brief background on islet vascular function and especially focuses on available techniques to measure islet blood perfusion. The gold standard for islet blood flow measurements in experimental animals is the microsphere technique, and its advantages and disadvantages will be discussed. In humans there are still no methods to measure islet blood flow selectively, but new developments in radiological techniques hold great hopes for the future. PMID:27124642

Fluid shear stress as a regulator of gene expression in vascular cells: possible correlations with diabetic abnormalities

NASA Technical Reports Server (NTRS)

Papadaki, M.; Eskin, S. G.; Ruef, J.; Runge, M. S.; McIntire, L. V.

1999-01-01

Diabetes mellitus is associated with increased frequency, severity and more rapid progression of cardiovascular diseases. Metabolic perturbations from hyperglycemia result in disturbed endothelium-dependent relaxation, activation of coagulation pathways, depressed fibrinolysis, and other abnormalities in vascular homeostasis. Atherosclerosis is localized mainly at areas of geometric irregularity at which blood vessels branch, curve and change diameter, and where blood is subjected to sudden changes in velocity and/or direction of flow. Shear stress resulting from blood flow is a well known modulator of vascular cell function. This paper presents what is currently known regarding the molecular mechanisms responsible for signal transduction and gene regulation in vascular cells exposed to shear stress. Considering the importance of the hemodynamic environment of vascular cells might be vital to increasing our understanding of diabetes.

[Survey of recent clinical trials of the prevention and immunointervention of type 1 diabetes mellitus].

PubMed

Boerschmann, H; Walter, M; Achenbach, P; Ziegler, A-G

2010-02-01

Immunomodulatory strategies in the management of type 1 diabetes mellitus (T1DM) have as their primary target the prevention of initiating islet autoimmunity (primary-), the secondary one is the progression to diabetes (secondary-) in non-diabetic persons at risk, and the decline of beta-cell function in new-onset patients (tertiary-prevention). This article reviews four recent immunointervention trials in patients with T1DM. (1) The Pre-POINT study is a primary prevention trial that will test whether vaccination with oral or nasal insulin can prevent the progression of islet autoimmunity and of T1DM in autoantibody-negative children who are genetically at high diabetes risk. (2) The Cord Blood study is a tertiary immunointervention trial that will test whether administration of autologous umbilical cord blood to children with T1DM can lead to regeneration of pancreatic islet insulin-producing beta-cells and improved blood glucose control. (3) The GAD Vaccination study will test whether vaccination with alum-formulated rhGAD65 (recombinant human glutamic acid decarboxylate) can preserve beta-cell function in 320 children with newly diagnosed T1DM, as has been suggested in a recent phase II study. (4) The AIDA study will test the beta-cell protective effect of interleukin-1-receptor antagonist Anakinra in 80 patients with T1DM, which has recently been shown to improve beta-cell function in patients with type 2 diabetes. Copyright Georg Thieme Verlag KG Stuttgart . New York.

BOLD delay times using group delay in sickle cell disease

NASA Astrophysics Data System (ADS)

Coloigner, Julie; Vu, Chau; Bush, Adam; Borzage, Matt; Rajagopalan, Vidya; Lepore, Natasha; Wood, John

2016-03-01

Sickle cell disease (SCD) is an inherited blood disorder that effects red blood cells, which can lead to vasoocclusion, ischemia and infarct. This disease often results in neurological damage and strokes, leading to morbidity and mortality. Functional Magnetic Resonance Imaging (fMRI) is a non-invasive technique for measuring and mapping the brain activity. Blood Oxygenation Level-Dependent (BOLD) signals contain also information about the neurovascular coupling, vascular reactivity, oxygenation and blood propagation. Temporal relationship between BOLD fluctuations in different parts of the brain provides also a mean to investigate the blood delay information. We used the induced desaturation as a label to profile transit times through different brain areas, reflecting oxygen utilization of tissue. In this study, we aimed to compare blood flow propagation delay times between these patients and healthy subjects in areas vascularized by anterior, middle and posterior cerebral arteries. In a group comparison analysis with control subjects, BOLD changes in these areas were found to be almost simultaneous and shorter in the SCD patients, because of their increased brain blood flow. Secondly, the analysis of a patient with a stenosis on the anterior cerebral artery indicated that signal of the area vascularized by this artery lagged the MCA signal. These findings suggest that sickle cell disease causes blood propagation modifications, and that these changes could be used as a biomarker of vascular damage.

Haematopoietic transplants combining a single unrelated cord blood unit and mobilized haematopoietic stem cells from an adult HLA-mismatched third party donor. Comparable results to transplants from HLA-identical related donors in adults with acute leukaemia and myelodysplastic syndromes.

PubMed

Sebrango, Ana; Vicuña, Isabel; de Laiglesia, Almudena; Millán, Isabel; Bautista, Guiomar; Martín-Donaire, Trinidad; Regidor, Carmen; Cabrera, Rafael; Fernandez, Manuel N

2010-06-01

We describe results of the strategy, developed by our group, of co-infusion of mobilized haematopoietic stem cells as a support for single-unit unrelated cord blood transplant (dual CB/TPD-MHSC transplants) for treatment of haematological malignancies in adults, and a comparative analysis of results obtained using this strategy and transplants performed with mobilized haematopoietic stem cells from related HLA-identical donors (RTD) for treatment of adults with acute leukaemia and myelodysplastic syndromes. Our data show that the dual CB/TPD-MHSC transplant strategy results in periods of post-transplant neutropenia, final rates of full donor chimerism and transplant-related mortality rates comparable to those of the RTD. Final survival outcomes are comparable in adults transplanted because of acute leukaemia, with different incidences of the complications that most influence these: a higher incidence of infections related to late recovery of protective immunity dependent on T cell functions, and a lower incidence of serious acute graft-versus-host disease and relapses. Recent advances in cord blood transplant techniques allow allogeneic haematopoietic stem cell transplantation (HSCT) to be a viable option for almost every patient who may benefit from this therapeutic approach. Development of innovative strategies to improve the post-transplant recovery of T cells function is currently the main challenge to further improving the possibilities of unrelated cord blood transplantation. Copyright © 2010 Elsevier Ltd. All rights reserved.

Compartmentalisation of innate immune responses in the central nervous system during cryptococcal meningitis/HIV co-infection

PubMed Central

NARANBHAI, Vivek; CHANG, Christina C.; DURGIAH, Raveshni; OMARJEE, Saleha; LIM, Andrew; MOOSA, Mahomed-Yunus S.; ELLIOT, Julian H.; NDUNG’U, Thumbi; LEWIN, Sharon R.; FRENCH, Martyn A.; CARR, William H.

2014-01-01

Objective The role of innate immunity in pathogenesis of cryptococcal meningitis (CM) is unclear. We hypothesised that NK cell and monocyte responses are central nervous system (CNS) compartmentalised, and altered by anti-fungal therapy and combination antiretroviral therapy (cART) during CM/HIV co-infection. Design Sub-study of a prospective cohort study of adults with CM/HIV co-infection in Durban, South Africa. Methods We used multi-parametric flow cytometry to study compartmentalisation of subsets, activation (CD69pos), CXCR3 and CX3CR1 expression and cytokine secretion of NK cells and monocytes in freshly collected blood and cerebrospinal fluid (CSF) at diagnosis (n=23), completion of anti-fungal therapy induction (n=19) and after a further 4 weeks of cART (n=9). Results Relative to blood, CSF was enriched with CD56bright (immunoregulatory) NK cells (p=0.0004). At enrolment, CXCR3 expression was more frequent amongst blood CD56bright than either blood CD56dim (p<0.0001) or CSF CD56bright (p=0.0002) NK cells. Anti-fungal therapy diminished blood (p<0.05) but not CSF CXCR3pos NK cell proportions nor CX3CR1pos NK cell proportions. CD56bright and CD56dim NK cells were more activated in CSF than blood (p<0.0001). Anti-fungal therapy induction reduced CD56dim NK cell activation in CSF (p=0.02). Activation of blood CD56bright and CD56dim NK cells was diminished following cART commencement (p<0.0001, p=0.03). Immunoregulatory NK cells in CSF tended to secrete higher levels of CXCL10 (p=0.06) and lower levels of TNF-α (p=0.06) than blood immunoregulatory NK cells. CSF was enriched with non-classical monocytes (p=0.001), but anti-fungal therapy restored proportions of classical monocytes (p=0.007). Conclusions These results highlight CNS activation, trafficking and function of NK cells and monocytes in CM/HIV and implicate immunoregulatory NK cells and pro-inflammatory monocytes as potential modulators of CM pathogenesis during HIV co-infection. PMID:24451162

Distribution of subpopulations of dendritic cells in peripheral blood of patients treated with exogenous thyrotropin

PubMed Central

2012-01-01

Background Dendritic cells (DCs) play a major role as regulators of inflammatory events associated with thyroid pathology. The immunoregulatory function of DCs depends strongly on their subtype, as well as maturation and activation status. Numerous hormonal factors modulate the immune properties of DCs, however, little is known about effects exerted by the hypothalamus-pituitary-thyroid-axis. Recently, we have shown a direct regulatory influence of thyroid hormones (TH) on human DCs function. The aim of the present study was to analyze the effect of systemically administered thyrotropin (TSH) on human blood DCs ex vivo. Methods Blood samples for the cytometric analysis of peripheral blood plasmacytoid and myeloid DCs subtypes were collected from patients subjected to total thyroidectomy because of differentiated thyroid carcinoma at 2 time points: (i) directly before the commencement of TSH administration and (ii) 5 days after first TSH injection. The whole blood quantitative and phenotypic analysis of plasmacytoid and myeloid DCs subtypes was performed by flow cytometry. Results Administration of TSH did not influence the percentage of plasmacytoid DCs in peripheral blood of study participants. Also the percentage of the two main myeloid DCs subpopulations – CD1c/BDCA1+ DCs and CD141/BDCA3+ DCs did not change significantly. TSH administration had no effect on the surface expression of CD86 – one of the major costimulatory molecules – neither in the whole peripheral blood mononuclear cell (PBMC) fraction nor in particular DCs subtypes. Conclusions In the present study, we demonstrated no influence of systemic TSH administration on human peripheral blood DCs subtypes. These results are in accordance with our previous work suggesting the direct effect of TH on human DCs ex vivo. PMID:23199104

Distribution of subpopulations of dendritic cells in peripheral blood of patients treated with exogenous thyrotropin.

PubMed

Stasiołek, Mariusz; Adamczewski, Zbigniew; Puła, Bartosz; Krawczyk-Rusiecka, Kinga; Zygmunt, Arkadiusz; Borowiecka, Magdalena; Dzięgiel, Piotr; Lewiński, Andrzej

2012-11-30

Dendritic cells (DCs) play a major role as regulators of inflammatory events associated with thyroid pathology. The immunoregulatory function of DCs depends strongly on their subtype, as well as maturation and activation status. Numerous hormonal factors modulate the immune properties of DCs, however, little is known about effects exerted by the hypothalamus-pituitary-thyroid-axis. Recently, we have shown a direct regulatory influence of thyroid hormones (TH) on human DCs function. The aim of the present study was to analyze the effect of systemically administered thyrotropin (TSH) on human blood DCs ex vivo. Blood samples for the cytometric analysis of peripheral blood plasmacytoid and myeloid DCs subtypes were collected from patients subjected to total thyroidectomy because of differentiated thyroid carcinoma at 2 time points: (i) directly before the commencement of TSH administration and (ii) 5 days after first TSH injection. The whole blood quantitative and phenotypic analysis of plasmacytoid and myeloid DCs subtypes was performed by flow cytometry. Administration of TSH did not influence the percentage of plasmacytoid DCs in peripheral blood of study participants. Also the percentage of the two main myeloid DCs subpopulations - CD1c/BDCA1+ DCs and CD141/BDCA3+ DCs did not change significantly. TSH administration had no effect on the surface expression of CD86 - one of the major costimulatory molecules - neither in the whole peripheral blood mononuclear cell (PBMC) fraction nor in particular DCs subtypes. In the present study, we demonstrated no influence of systemic TSH administration on human peripheral blood DCs subtypes. These results are in accordance with our previous work suggesting the direct effect of TH on human DCs ex vivo.

Progenitor cell dose determines the pace and completeness of engraftment in a xenograft model for cord blood transplantation

PubMed Central

Liu, Congxiao; Chen, Benny J.; DeOliveira, Divinomar; Sempowski, Gregory D.; Chao, Nelson J.

2010-01-01

Two critical concerns in clinical cord blood transplantation are the initial time to engraftment and the subsequent restoration of immune function. These studies measured the impact of progenitor cell dose on both the pace and strength of hematopoietic reconstitution by transplanting nonobese diabetic/severe combined immunodeficiency/interleukin-2 receptor-gamma–null (NSγ) mice with lineage-depleted aldehyde dehydrogenase-bright CD34+ human cord blood progenitors. The progress of each transplant was monitored over an extended time course by repeatedly analyzing the peripheral blood for human hematopoietic cells. In vivo human hematopoietic development was complete. After long-term transplantation assays (≥ 19 weeks), human T-cell development was documented within multiple tissues in 16 of 32 NSγ mice. Human T-cell differentiation was active within NSγ thymuses, as documented by the presence of CD4+ CD8+ T-cell progenitors as well as T-cell receptor excision circles. It is important to note that although myeloid and B-cell engraftment was detected as early as 4 weeks after transplantation, human T-cell development was exclusively late onset. High progenitor cell doses were associated with a robust human hematopoietic chimerism that accelerated both initial time to engraftment and subsequent T-cell development. At lower progenitor cell doses, the chimerism was weak and the human hematopoietic lineage development was frequently incomplete. PMID:20833978

The use of biomarkers to describe plasma-, red cell-, and blood volume from a simple blood test.

PubMed

Lobigs, Louisa Margit; Sottas, Pierre-Edouard; Bourdon, Pitre Collier; Nikolovski, Zoran; El-Gingo, Mohamed; Varamenti, Evdokia; Peeling, Peter; Dawson, Brian; Schumacher, Yorck Olaf

2017-01-01

Plasma volume and red cell mass are key health markers used to monitor numerous disease states, such as heart failure, kidney disease, or sepsis. Nevertheless, there is currently no practically applicable method to easily measure absolute plasma or red cell volumes in a clinical setting. Here, a novel marker for plasma volume and red cell mass was developed through analysis of the observed variability caused by plasma volume shifts in common biochemical measures, selected based on their propensity to present with low variations over time. Once a month for 6 months, serum and whole blood samples were collected from 33 active males. Concurrently, the CO-rebreathing method was applied to determine target levels of hemoglobin mass (HbM) and blood volumes. The variability of 18 common chemistry markers and 27 Full Blood Count variables was investigated and matched to the observed plasma volume variation. After the removal of between-subject variations using a Bayesian model, multivariate analysis identified two sets of 8 and 15 biomarkers explaining 68% and 69% of plasma volume variance, respectively. The final multiparametric model contains a weighting function to allow for isolated abnormalities in single biomarkers. This proof-of-concept investigation describes a novel approach to estimate absolute vascular volumes, with a simple blood test. Despite the physiological instability of critically ill patients, it is hypothesized the model, with its multiparametric approach and weighting function, maintains the capacity to describe vascular volumes. This model has potential to transform volume management in clinical settings. Am. J. Hematol. 92:62-67, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

T-cell phenotypes in bronchoalveolar lavage fluid, blood and lymph nodes in pulmonary sarcoidosis--indication for an airborne antigen as the triggering factor in sarcoidosis.

PubMed

Darlington, P; Haugom-Olsen, H; von Sivers, K; Wahlström, J; Runold, M; Svjatoha, V; Porwit, A; Eklund, A; Grunewald, J

2012-11-01

An increased percentage of CD4+ T cells is usually observed in bronchoalveolar lavage fluid (BALF) from patients with sarcoidosis. In HLA-DRB1*03-positive patients, such T cells express the T-cell receptor (TCR) AV2S3+ gene segment. It is not known whether cells found in BALF reflect those in enlarged regional lymph nodes (LNs). Therefore, the aim of this study was to compare T-cell phenotypes in BALF, blood and mediastinal LNs. Fifteen patients underwent clinical investigation including bronchoscopy with bronchoalveolar lavage. Blood samples were drawn, and endoscopic ultrasound-guided fine-needle aspiration of enlarged mediastinal LNs was performed via the oesophagus. T cells from all three compartments were analysed by flow cytometry for markers of activity, differentiation and T regulatory function. The CD4/CD8 ratio was significantly higher in BALF compared with regional LNs and was also significantly higher in LNs than in blood. The CD4+ T cells were recently activated and more differentiated in BALF than in blood and LNs. There was an accumulation of T regulatory cells (FOXP3+) in LNs and a correlation between high levels of FOXP3+ cells in BALF and in LNs. In HLA-DRB1*03-positive patients, TCR AV2S3+ CD4+ T cells were predominantly localized within BALF. The CD4+ T-cell phenotype in BALF indicates an active ongoing specific immune response primarily localized to the alveolar space. © 2012 The Association for the Publication of the Journal of Internal Medicine.

Whole Blood Activation Results in Altered T Cell and Monocyte Cytokine Production Profiles by Flow Cytometry

NASA Technical Reports Server (NTRS)

Crucian, Brian E.; Sams, Clarence F.

2001-01-01

An excellent monitor of the immune balance of peripheral circulating cells is to determine their cytokine production patterns in response to stimuli. Using flow cytometry, a positive identification of cytokine producing cells in a mixed culture may be achieved. Recently, the ability to assess cytokine production following a whole-blood activation culture has been described. In this study, whole blood activation was compared to traditional PBMC activation and the individual cytokine secretion patterns for both T cells, T cell subsets and monocytes was determined by flow cytometry. RESULTS: For T cell cytokine assessment (IFNg/IL-10 and IL-21/L-4) following PMA +ionomycin activation: (1) a Significantly greater percentages of T cells producing IFNgamma and IL-2 were observed following whole-blood culture and (2) altered T cell cytokine production kinetics were observed by varying whole blood culture times. Four-color analysiS was used to allow assessment of cytokine production by specific T cell subsets. It was found that IFNgamma production was significantly elevated in the CD3+/CD8+ T cell population as compared to the CD3+/CD8- population following five hours of whole blood activation. Conversely, IL-2 and IL-10 production were Significantly elevated in the CD3+/CD8- T cell population as compared to the CD3+/CD8+ population. Monocyte cytokine production was assessed in both culture systems following LPS activation for 24 hours. A three-color flow cytometric was used to assess two cytokines (IL-1a/IL-12 and TNFa/IL-10) in conjunction with CD14. Nearly all monocytes were stimulated to produce IL-1a, IL-12 and TNFa. equally well in both culture systems, however monocyte production of IL-10 was significantly elevated in whole blood culture as compared to PBMC culture. IL-12 producing monocytes appeared to be a distinct subpopulation of the IL-1a producing set, whereas IL-10 and TNFa producing monocytes were largely mutually exclusive. IL-10 and TNFa producing monocytes may represent distinct monocyte subsets with unique functions. CONCLUSIONS: Whole blood culture eliminates the need to purify cell populations prior to culture and may have Significant utility for the routine monitoring of the cytokine balances of the peripheral blood T cell and monocyte populations. In addition, there are distinct advantages to performing whole-blood (WB) activation as compared to PBMC activation. These advantages would include retaining all various cell-cell interactions as well as any soluble factors present in serum that influence cell activation. In this study, alterations in cytokine production are demonstrated between whole blood and PBMC activation. It is likely that whole blood activation more accurately represents the in-vivo immune balance than PBMC activation.

Regulation of hematopoietic stem cell aging by the small RhoGTPase Cdc42

PubMed Central

Geiger, Hartmut; Zheng, Yi

2015-01-01

Summary Aging of stem cells might be the underlying cause of tissue aging in tissue that in the adult heavily rely on stem cell activity, like the blood forming system. Hematopoiesis, the generation of blood forming cells, is sustained by hematopoietic stem cells. In this review article, we introduce the canonical set of phenotypes associated with aged HSCs, focus on the novel aging-associated phenotype apolarity caused by elevated activity of the small RhoGTPase in aged HSCs, disuccs the role of Cdc42 in hematopoiesis and describe that pharmacological inhibition of Cdc42 activity in aged HSCs results in functionally young and thus rejuvenated HSCs. PMID:25220425

Functional and phenotypic characterization of CD8+CD28+ and CD28- T cells in atopic individuals sensitized to Dermatophagoides pteronyssinus.

PubMed

Lourenço, O; Fonseca, A M; Paiva, A; Arosa, F A; Taborda-Barata, L

2006-01-01

CD8+ T suppressor cells may play a role in immunoregulation. Recent studies have characterized this population by the lack of the CD28 molecule. These CD8+CD28 T cells differ phenotypically and functionally from CD8 + CD28 + T cells. Little is known about CD8 + CD28 cells in atopy. Our aim was to analyze the phenotype and functional properties of CD8 + CD28T cells in atopic and non-atopic individuals. Peripheral blood mononuclear cells (PBMC) were obtained after density gradient centrifugation. CD8 + CD28 and CD8 + CD28 + T cells were isolated using immunomagnetic beads. Relative percentages of these cells and expression of several phenotypic markers were analyzed by flow cytometry. Proliferation was assessed by thymidine incorporation in isolated populations and in co-cultures with PBMC using Dermatophagoides pteronyssinus as stimulus. Cytokine synthesis was evaluated in culture supernatants by cytometric bead array. The relative percentages of CD8+CD28 T cells and their phenotypic expression in atopic and non-atopic volunteers were not significantly different. However, CD8 + CD28 T cells showed greater proliferation than did CD8+CD28+ T cells when stimulated with D. pteronyssinus, although cytokine synthesis patterns were similar. CD8+CD28 co-cultures with PBMC showed greater proliferation than CD8+CD28+ T cell co-cultures, but cytokine synthesis patterns were not different. Our data confirm phenotypic and functional differences between CD28+ and CD28 T cells, irrespective of atopic status. Purified human CD8+CD28 T cells, freshly isolated from peripheral blood, do not have suppressor properties on allergen-specific proliferation or on cytokine synthesis in PBMC.

Red blood cells as modulators of T cell growth and survival.

PubMed

Arosa, Fernando A; Pereira, Carlos F; Fonseca, Ana M

2004-01-01

T cell homeostasis is largely controlled by a balance between cell death and survival and anomalies in either process account for a number of diseases linked to excessive or faulty T cell growth. Yet, the influence that cells outside the immunological system have on these processes has only recently received attention. Accumulated evidence indicate that homeostasis of the CD4+ and CD8+ T cell pools is highly dynamic and regulated by signals delivered by cells and molecules present in the different internal microenvironments. The major function of red blood cells (RBC) is generally considered to be oxygen and carbon dioxide transport. In recent years, however, RBC have been implicated in the regulation of basic physiological processes, from vascular contraction and platelet aggregation to T cell growth and survival. Regulation of T cell survival by RBC may influence the response of selected subsets of T cells to internal or external stimuli and may help explaining the immunomodulatory activities of red blood cells. By interfering in the balance between death and survival RBC become potential tools that can be manipulated to improve or reverse pathological situations characterized by anomalies in the control of T cell growth.

Characterization of Tumor Cells Using a Medical Wire for Capturing Circulating Tumor Cells: A 3D Approach Based on Immunofluorescence and DNA FISH

PubMed Central

Gallerani, Giulia; Cocchi, Claudia; Bocchini, Martine; Piccinini, Filippo; Fabbri, Francesco

2017-01-01

Circulating tumor cells (CTCs) are associated with poor survival in metastatic cancer. Their identification, phenotyping, and genotyping could lead to a better understanding of tumor heterogeneity and thus facilitate the selection of patients for personalized treatment. However, this is hampered because of the rarity of CTCs. We present an innovative approach for sampling a high volume of the patient blood and obtaining information about presence, phenotype, and gene translocation of CTCs. The method combines immunofluorescence staining and DNA fluorescent-in-situ-hybridization (DNA FISH) and is based on a functionalized medical wire. This wire is an innovative device that permits the in vivo isolation of CTCs from a large volume of peripheral blood. The blood volume screened by a 30-min administration of the wire is approximately 1.5-3 L. To demonstrate the feasibility of this approach, epithelial cell adhesion molecule (EpCAM) expression and the chromosomal translocation of the ALK gene were determined in non-small-cell lung cancer (NSCLC) cell lines captured by the functionalized wire and stained with an immuno-DNA FISH approach. Our main challenge was to perform the assay on a 3D structure, the functionalized wire, and to determine immuno-phenotype and FISH signals on this support using a conventional fluorescence microscope. The results obtained indicate that catching CTCs and analyzing their phenotype and chromosomal rearrangement could potentially represent a new companion diagnostic approach and provide an innovative strategy for improving personalized cancer treatments. PMID:29286485

Identification of Differentially Expressed Genes in Blood Cells of Narcolepsy Patients

PubMed Central

Tanaka, Susumu; Honda, Yutaka; Honda, Makoto

2007-01-01

Study Objective: A close association between the human leukocyte antigen (HLA)-DRB1*1501/DQB1*0602 and abnormalities in some inflammatory cytokines have been demonstrated in narcolepsy. Specific alterations in the immune system have been suggested to occur in this disorder. We attempted to identify alterations in gene expression underlying the abnormalities in the blood cells of narcoleptic patients. Designs: Total RNA from 12 narcolepsy-cataplexy patients and from 12 age- and sex-matched healthy controls were pooled. The pooled samples were initially screened for candidate genes for narcolepsy by differential display analysis using annealing control primers (ACP). The second screening of the samples was carried out by semiquantitative PCR using gene-specific primers. Finally, the expression levels of the candidate genes were further confirmed by quantitative real-time PCR using a new set of samples (20 narcolepsy-cataplexy patients and 20 healthy controls). Results: The second screening revealed differential expression of 4 candidate genes. Among them, MX2 was confirmed as a significantly down-regulated gene in the white blood cells of narcoleptic patients by quantitative real-time PCR. Conclusion: We found the MX2 gene to be significantly less expressed in comparison with normal subjects in the white blood cells of narcoleptic patients. This gene is relevant to the immune system. Although differential display analysis using ACP technology has a limitation in that it does not help in determining the functional mechanism underlying sleep/wakefulness dysregulation, it is useful for identifying novel genetic factors related to narcolepsy, such as HLA molecules. Further studies are required to explore the functional relationship between the MX2 gene and narcolepsy pathophysiology. Citation: Tanaka S; Honda Y; Honda M. Identification of differentially expressed genes in blood cells of narcolepsy patients. SLEEP 2007;30(8):974-979. PMID:17702266

Health care resource utilization and characteristics of patients with eosinophilic asthma in secondary health care in Finland.

PubMed

Mäkelä, Mika J; Christensen, Helene Nordahl; Karlsson, Antti; Rastogi, Sarang; Kettunen, Kirsi

2018-01-01

Background : Eosinophilic airway inflammation is common in asthma patients and appears to be associated with severe exacerbations and loss of asthma control. Objective : To describe the resource utilization and clinical characteristics of patients with eosinophilic asthma. Design : Asthma patients ≥18 years with ≥1 blood eosinophil count in secondary care (South West Finland) during 2003‒2013 were included. Clinical characteristics (age, lung function, body mass index, and comorbidities) and asthma-related resource utilization (hospital admissions, outpatient visits, and emergency room [ER] visits) were retrieved. Resource utilization rates were compared for patients with blood eosinophil ≤ or >300 cells/μL, using adjusted negative binomial regression models. Results : Overall, 4,357 eligible patients were identified (mean age 60 years, females 68%), of which 1,927 (44%) had >300 eosinophil cells/μL blood. Patients with ≤300 and >300 eosinophil counts, exhibited similar clinical characteristics, including advanced age, poor lung function, and overweight. Comorbidities such as pneumonia, sinusitis, and nasal polyps, were more frequent among those with >300 eosinophil cells/μL blood compared with patients with lower counts. Eosinophil counts >300 cells/μL were associated with greater hospital admissions (rate ratio [RR] [95% confidence interval CI]: 1.13 [1.02;1.24]) and outpatient visits (RR [95% CI]: 1.11 [1.03;1.20]) compared with patients with lower eosinophil counts. Rates of ER visits were similar between the patient groups (RR [95% CI]: 0.99 [0.87;1.12]). Conclusions : Hospital admissions and outpatient visits occurred more often for patients with eosinophil counts >300 cells/µL, than for patients with lower eosinophil counts. Routine blood eosinophil screening might be useful to identify patients with an eosinophilic phenotype eligible for more targeted treatments.

Lymphatic endothelial cell line (CH3) from a recurrent retroperitoneal lymphangioma.

PubMed

Way, D; Hendrix, M; Witte, M; Witte, C; Nagle, R; Davis, J

1987-09-01

An endothelial cell line derived from a massive recurrent chyle-containing retroperitoneal lymphangioma was isolated in monolayer culture. Scanning and transmission electron microscopy and immunohistochemistry confirmed a close resemblance to blood vascular endothelium with typical cobblestone morphology, positive immunofluorescence staining for endothelial marker Factor VIII-associated antigen and fibronectin, and prominent Weibel-Palade bodies. The endothelial cells also exhibited other ultrastructural features characteristic of lymphatic endothelium, including sparse microvillous surface projections, overlapping intercellular junctions, and abundant intermediate filaments. This endothelial cell line represents a new source of proliferating lymphatic endothelium for future study, including structural and functional comparison to blood vascular endothelium.

Oxygen transport in blood at high altitude: role of the hemoglobin-oxygen affinity and impact of the phenomena related to hemoglobin allosterism and red cell function.

PubMed

Samaja, Michele; Crespi, Tiziano; Guazzi, Marco; Vandegriff, Kim D

2003-10-01

Altitude hypoxia is a major challenge to the blood O2 transport system, and adjustments of the blood-O2 affinity might contribute significantly to hypoxia adaptation. In principle, lowering the blood-O2 affinity is advantageous because it lowers the circulatory load required to assure adequate tissue oxygenation up to a threshold corresponding to about 5,000 m altitude, whereas at higher altitudes an increased blood-O2 affinity appears more advantageous. However, the rather contradictory experimental evidence raises the question whether other factors superimpose on the apparent changes of the blood-O2 affinity. The most important of these are as follows: (1) absolute temperature and temperature gradients within the body; (2) the intracapillary Bohr effect; (3) the red cell population heterogeneity in terms of O2 affinity; (4) control of altitude alkalosis; (5) the possible role of hemoglobin as a carrier of the vasodilator nitric oxide; (6) the effect of varied red cell transit times through the capillaries.

Low Immunogenic Endothelial Cells Maintain Morphological and Functional Properties Required for Vascular Tissue Engineering.

PubMed

Lau, Skadi; Eicke, Dorothee; Carvalho Oliveira, Marco; Wiegmann, Bettina; Schrimpf, Claudia; Haverich, Axel; Blasczyk, Rainer; Wilhelmi, Mathias; Figueiredo, Constança; Böer, Ulrike

2018-03-01

The limited availability of native vessels suitable for the application as hemodialysis shunts or bypass material demands new strategies in cardiovascular surgery. Tissue-engineered vascular grafts containing autologous cells are considered ideal vessel replacements due to the low risk of rejection. However, endothelial cells (EC), which are central components of natural blood vessels, are difficult to obtain from elderly patients of poor health. Umbilical cord blood represents a promising alternative source for EC, but their allogeneic origin corresponds with the risk of rejection after allotransplantation. To reduce this risk, the human leukocyte antigen class I (HLA I) complex was stably silenced by lentiviral vector-mediated RNA interference (RNAi) in EC from peripheral blood and umbilical cord blood and vein. EC from all three sources were transduced by 93.1% ± 4.8% and effectively, HLA I-silenced by up to 67% compared to nontransduced (NT) cells or transduced with a nonspecific short hairpin RNA, respectively. Silenced EC remained capable to express characteristic endothelial surface markers such as CD31 and vascular endothelial cadherin important for constructing a tight barrier, as well as von Willebrand factor and endothelial nitric oxide synthase important for blood coagulation and vessel tone regulation. Moreover, HLA I-silenced EC were still able to align under unidirectional flow, to take up acetylated low-density lipoprotein, and to form capillary-like tube structures in three-dimensional fibrin gels similar to NT cells. In particular, addition of adipose tissue-derived mesenchymal stem cells significantly improved tube formation capability of HLA I-silenced EC toward long and widely branched vascular networks necessary for prevascularizing vascular grafts. Thus, silencing HLA I by RNAi represents a promising technique to reduce the immunogenic potential of EC from three different sources without interfering with EC-specific morphological and functional properties required for vascular tissue engineering. This extends the spectrum of available cell sources from autologous to allogeneic sources, thereby accelerating the generation of tissue-engineered vascular grafts in acute clinical cases.

Identification of stem cells from human umbilical cord blood with embryonic and hematopoietic characteristics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao Yong; Wang Honglan; Mazzone, Theodore

2006-08-01

We identified stem cells from the umbilical cord blood, designated cord blood-stem cells (CB-SC). CB-SC displayed important embryonic stem (ES) cell characteristics including expression of ES-cell-specific molecular markers including transcription factors OCT-4 and Nanog, along with stage-specific embryonic antigen (SSEA)-3 and SSEA-4. CB-SC also expressed hematopoietic cell antigens including CD9, CD45 and CD117, but were negative for CD34. CB-SC displayed very low immunogenicity as indicated by expression of a very low level of major histocompatibility complex (MHC) antigens and failure to stimulate the proliferation of allogeneic lymphocytes. CB-SC could give rise to cells with endothelial-like and neuronal-like characteristics in vitro,more » as demonstrated by expression of lineage-associated markers. Notably, CB-SC could be stimulated to differentiate into functional insulin-producing cells in vivo and eliminated hyperglycemia after transplantation into a streptozotocin-induced diabetic mouse model. These findings may have significant potential to advance stem-cell-based therapeutics.« less

Human Naive T Cells Express Functional CXCL8 and Promote Tumorigenesis.

PubMed

Crespo, Joel; Wu, Ke; Li, Wei; Kryczek, Ilona; Maj, Tomasz; Vatan, Linda; Wei, Shuang; Opipari, Anthony W; Zou, Weiping

2018-05-25

Naive T cells are thought to be functionally quiescent. In this study, we studied and compared the phenotype, cytokine profile, and potential function of human naive CD4 + T cells in umbilical cord and peripheral blood. We found that naive CD4 + T cells, but not memory T cells, expressed high levels of chemokine CXCL8. CXCL8 + naive T cells were preferentially enriched CD31 + T cells and did not express T cell activation markers or typical Th effector cytokines, including IFN-γ, IL-4, IL-17, and IL-22. In addition, upon activation, naive T cells retained high levels of CXCL8 expression. Furthermore, we showed that naive T cell-derived CXCL8 mediated neutrophil migration in the in vitro migration assay, supported tumor sphere formation, and promoted tumor growth in an in vivo human xenograft model. Thus, human naive T cells are phenotypically and functionally heterogeneous and can carry out active functions in immune responses. Copyright © 2018 by The American Association of Immunologists, Inc.

Structural, functional and blood perfusion changes in the rat retina associated with elevated intraocular pressure, measured simultaneously with a combined OCT+ERG system

PubMed Central

Tan, Bingyao; MacLellan, Benjamin; Mason, Erik

2018-01-01

Acute elevation of intraocular pressure (IOP) to ischemic and non-ischemic levels can cause temporary or permanent changes in the retinal morphology, function and blood flow/blood perfusion. Previously, such changes in the retina were assessed separately with different methods in clinical studies and animal models. In this study, we used a combined OCT+ ERG system in combination with Doppler OCT and OCT angiography (OCTA) imaging protocols, in order to evaluate simultaneously and correlate changes in the retinal morphology, the retinal functional response to visual stimulation, and the retinal blood flow/blood perfusion, associated with IOP elevation to ischemic and non-ischemic levels in rats. Results from this study suggest that the inner retina responds faster to IOP elevation to levels greater than 30 mmHg with significant reduction of the total retinal blood flow (TRBF), decrease of the capillaries’ perfusion and reduction of the ON bipolar cells contribution to the ERG traces. Furthermore, this study showed that ischemic levels of IOP elevation cause an additional significant decrease in the ERG photoreceptor response in the posterior retina. Thirty minutes after IOP normalization, retinal morphology, blood flow and blood perfusion recovered to baseline values, while retinal function did not recover completely. PMID:29509807

Relationship of Blood Mercury Levels to Health Parameters in the Loggerhead Sea Turtle (Caretta caretta)

PubMed Central

Day, Rusty D.; Segars, Al L.; Arendt, Michael D.; Lee, A. Michelle; Peden-Adams, Margie M.

2007-01-01

Background Mercury is a pervasive environmental pollutant whose toxic effects have not been studied in sea turtles in spite of their threatened status and evidence of immunosuppression in diseased populations. Objectives In the present study we investigate mercury toxicity in loggerhead sea turtles (Caretta caretta) by examining trends between blood mercury concentrations and various health parameters. Methods Blood was collected from free-ranging turtles, and correlations between blood mercury concentrations and plasma chemistries, complete blood counts, lysozyme, and lymphocyte proliferation were examined. Lymphocytes were also harvested from free-ranging turtles and exposed in vitro to methylmercury to assess proliferative responses. Results Blood mercury concentrations were positively correlated with hematocrit and creatine phosphokinase activity, and negatively correlated with lymphocyte cell counts and aspartate amino-transferase. Ex vivo negative correlations between blood mercury concentrations and B-cell proliferation were observed in 2001 and 2003 under optimal assay conditions. In vitro exposure of peripheral blood leukocytes to methylmercury resulted in suppression of proliferative responses for B cells (0.1 μg/g and 0.35 μg/g) and T cells (0.7 μg/g). Conclusions The positive correlation between blood mercury concentration and hematocrit reflects the higher affinity of mercury species for erythrocytes than plasma, and demonstrates the importance of measuring hematocrit when analyzing whole blood for mercury. In vitro immunosuppression occurred at methylmercury concentrations that correspond to approximately 5% of the individuals captured in the wild. This observation and the negative correlation found ex vivo between mercury and lymphocyte numbers and mercury and B-cell proliferative responses suggests that subtle negative impacts of mercury on sea turtle immune function are possible at concentrations observed in the wild. PMID:17938730

Elastomeric negative acoustic contrast particles for affinity capture assays.

PubMed

Cushing, Kevin W; Piyasena, Menake E; Carroll, Nick J; Maestas, Gian C; López, Beth Ann; Edwards, Bruce S; Graves, Steven W; López, Gabriel P

2013-02-19

This report describes the development of elastomeric capture microparticles (ECμPs) and their use with acoustophoretic separation to perform microparticle assays via flow cytometry.We have developed simple methods to form ECμPs by cross-linking droplets of common commercially available silicone precursors in suspension followed by surface functionalization with biomolecular recognition reagents. The ECμPs are compressible particles that exhibit negative acoustic contrast in ultrasound when suspended in aqueous media, blood serum, or diluted blood. In this study, these particles have been functionalized with antibodies to bind prostate specific antigen and immunoglobulin (IgG). Specific separation of the ECμPs from blood cells is achieved by flowing them through a microfluidic acoustophoretic device that uses an ultrasonic standing wave to align the blood cells, which exhibit positive acoustic contrast, at a node in the acoustic pressure distribution while aligning the negative acoustic contrast ECμPs at the antinodes. Laminar flow of the separated particles to downstream collection ports allows for collection of the separated negative contrast (ECμPs) and positive contrast particles (cells). Separated ECμPs were analyzed via flow cytometry to demonstrate nanomolar detection for prostate specific antigen in aqueous buffer and picomolar detection for IgG in plasma and diluted blood samples. This approach has potential applications in the development of rapid assays that detect the presence of low concentrations of biomarkers in a number of biological sample types.

Elastomeric Negative Acoustic Contrast Particles for Affinity Capture Assays

PubMed Central

Cushing, Kevin W.; Piyasena, Menake E.; Carroll, Nick J.; Maestas, Gian C.; López, Beth Ann; Edwards, Bruce S.; Graves, Steven W.; López, Gabriel P.

2013-01-01

This report describes the development of elastomeric capture microparticles (ECμPs) and their use with acoustophoretic separation to perform microparticle assays via flow cytometry. We have developed simple methods to form ECμPsby crosslinking droplets of common commercially available silicone precursors in suspension followed by surface functionalization with biomolecular recognition reagents. The ECμPs are compressible particles that exhibit negative acoustic contrast in ultrasound when suspended in aqueous media, blood serum or diluted blood. In this study, these particles have been functionalized with antibodies to bind prostate specific antigen and immunoglobulin (IgG). Specific separation of the ECμPs from blood cells is achieved by flowing them through a microfluidic acoustophoretic device that uses an ultrasonic standing wave to align the blood cells, which exhibit positive acoustic contrast, at a node in the acoustic pressure distribution while aligning the negative acoustic contrast ECμPs at the antinodes. Laminar flow of the separated particles to downstream collection ports allows for collection of the separated negative contrast (ECμPs) and positive contrast particles (cells). Separated ECμPs were analyzed via flow cytometry to demonstrate nanomolar detection for prostate specific antigen in aqueous buffer and picomolar detection for IgG in plasma and diluted blood samples. This approach has potential applications in the development of rapid assays that detect the presence of low concentrations of biomarkers in a number of biological sample types. PMID:23331264

DECREASED LEVEL OF CORD BLOOD CIRCULATING ENDOTHELIAL COLONY-FORMING CELLS IN PREECLAMPSIA

PubMed Central

Muñoz-Hernandez, Rocio; Miranda, Maria L.; Stiefel, Pablo; Lin, Ruei-Zeng; Praena-Fernández, Juan M.; Dominguez-Simeon, Maria J.; Villar, Jose; Moreno-Luna, Rafael; Melero-Martin, Juan M.

2014-01-01

Preeclampsia is a pregnancy-related disorder associated with increased cardiovascular risk for the offspring. Endothelial colony-forming cells (ECFCs) are a subset of circulating endothelial progenitor cells that participate in the formation of vasculature during development. However, the effect of preeclampsia on fetal levels of ECFCs is largely unknown. In this study, we sought to determine whether cord blood ECFC abundance and function are altered in preeclampsia. We conducted a prospective cohort study that included women with normal (n=35) and preeclamptic (n=15) pregnancies. We measured ECFC levels in the umbilical cord blood of neonates and characterized ECFC phenotype, cloning-forming ability, proliferation and migration towards VEGF-A and FGF-2, in vitro formation of capillary-like structures, and in vivo vasculogenic ability in immunodeficient mice. We found that the level of cord blood ECFCs was statistically lower in preeclampsia than in control pregnancies (P = .04), a reduction that was independent of other obstetric factors. In addition, cord blood ECFCs from preeclamptic pregnancies required more time to emerge in culture than control ECFCs. However, once derived in culture, ECFC function was deemed normal and highly similar between preeclampsia and control, including the ability to form vascular networks in vivo. This study demonstrates that preeclampsia affects ECFC abundance in neonates. A reduced level of ECFCs during preeclamptic pregnancies may contribute to an increased risk of developing future cardiovascular events. PMID:24752434

Reconstructing blood stem cell regulatory network models from single-cell molecular profiles

PubMed Central

Hamey, Fiona K.; Nestorowa, Sonia; Kinston, Sarah J.; Kent, David G.; Wilson, Nicola K.

2017-01-01

Adult blood contains a mixture of mature cell types, each with specialized functions. Single hematopoietic stem cells (HSCs) have been functionally shown to generate all mature cell types for the lifetime of the organism. Differentiation of HSCs toward alternative lineages must be balanced at the population level by the fate decisions made by individual cells. Transcription factors play a key role in regulating these decisions and operate within organized regulatory programs that can be modeled as transcriptional regulatory networks. As dysregulation of single HSC fate decisions is linked to fatal malignancies such as leukemia, it is important to understand how these decisions are controlled on a cell-by-cell basis. Here we developed and applied a network inference method, exploiting the ability to infer dynamic information from single-cell snapshot expression data based on expression profiles of 48 genes in 2,167 blood stem and progenitor cells. This approach allowed us to infer transcriptional regulatory network models that recapitulated differentiation of HSCs into progenitor cell types, focusing on trajectories toward megakaryocyte–erythrocyte progenitors and lymphoid-primed multipotent progenitors. By comparing these two models, we identified and subsequently experimentally validated a difference in the regulation of nuclear factor, erythroid 2 (Nfe2) and core-binding factor, runt domain, alpha subunit 2, translocated to, 3 homolog (Cbfa2t3h) by the transcription factor Gata2. Our approach confirms known aspects of hematopoiesis, provides hypotheses about regulation of HSC differentiation, and is widely applicable to other hierarchical biological systems to uncover regulatory relationships. PMID:28584094

Mesenchymal stem cell and derived exosome as small RNA carrier and Immunomodulator to improve islet transplantation.

PubMed

Wen, Di; Peng, Yang; Liu, Di; Weizmann, Yossi; Mahato, Ram I

2016-09-28

Human bone marrow mesenchymal stem cells (hBMSCs) and their exosomes can suppress immune reaction and deliver small RNAs. Thus, they may improve islet transplantation by delivering small RNAs for promoting islet function and inhibiting immune rejection. Here, we proposed an hBMSC and its exosome-based therapy to overcome immune rejection and poor islet function, both of which hinder the success of islet transplantation. We found overexpressed siFas and anti-miR-375 in plasmid encoding shFas and anti-miR-375 transfected hBMSC-derived exosomes, which silenced Fas and miR-375 of human islets and improved their viability and function against inflammatory cytokines. This plasmid transfected hBMSCs downregulated Fas and miR-375 of human islets in a humanized NOD scid gamma (NSG) mouse model, whose immune reaction was inhibited by injecting hBMSC and peripheral blood mononuclear cell (PBMC) co-cultured exosomes. These exosomes suppressed immune reaction by inhibiting PBMC proliferation and enhancing regulatory T cell (Treg) function. Collectively, our studies elucidated the mechanisms of RNA delivery from hBMSCs to human islets and the immunosuppressive effect of hBMSC and peripheral blood mononuclear cell co-cultured exosomes for improving islet transplantation. Copyright © 2016 Elsevier B.V. All rights reserved.

Effects of alcohols and ciclopirox alamine, an anti-fungal agent, on the peripheral blood lymphocyte functions in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Prabhala, R.H.; Maxey, V.; Hicks, M.J.

1986-03-01

Effects of ethanol (1%), propanol (1%) and ciclopirox alamine, an anti-fungal agent, (4 ..mu..g/well), on the peripheral blood lymphocyte functions, including response to T- (Concanavalin A, ConA) and B-cell (Lipopolysaccharide, LPS) mitogens, and presence of functional T-lymphocyte subsets were determined in vitro. Purified human lymphocytes were incubated at 37/sup 0/C for 48 hours with or without test compounds in presence or absence of ConA and LPS. All three compounds suppress the response to T- or B-cell mitogens. The percentage of T-lymphocytes with T-helper characteristics in the presence of ethanol and ciclopirox alamine was increased. All three compounds suppressed the percentagemore » of T-lymphocytes with E-resetting characteristics. Alcohols enhanced the number of natural killer cells, whereas, ciclopirox alamine exhibited the reverse action. Although the alcohols and the anti-fungal agent enhanced the T-helper subpopulation, their response to mitogens was suppressed. This may be due to the suppression of T-cell activating lymphokines. Alcohol metabolite such as acetaldehyde also suppress the number of T-cells and their functions at 0.01% so may also be part of the explanation for immunoalteration.« less

Effects of Fenbendazole on Routine Immune Response Parameters of BALB/c Mice

PubMed Central

Cray, Carolyn; Villar, David; Zaias, Julia; Altman, Norman H

2008-01-01

Fenbendazole (FBZ) is an anthelmintic drug widely used to treat and prevent pinworm outbreaks in laboratory rodents. Although data in nonrodent species indicate possible effects of fenbendazole on the bone marrow and lymphocyte proliferation and function, little has been reported regarding possible effects on the rodent immune system. The purpose of the current study was to determine the effects of a therapeutic regimen of FBZ on immune parameters in BALB/c mice. Both 9-wk on–off and 5-wk continuous medicated feed protocols were assessed. No significant differences between normal and FBZ diet treated mice were observed in the following parameters: complete blood count, blood chemistry, quantitation of major T and B cell markers in spleen, quantitation of T cell markers in the thymus, spleen cell proliferation to T and B cell mitogens, bone marrow colony-forming cell assays, skin graft rejection, and primary and secondary humoral immune responses. These data indicate that FBZ treatment does not affect many standard broad measures of immune function. PMID:19049250

Effects of fenbendazole on routine immune response parameters of BALB/c mice.

PubMed

Cray, Carolyn; Villar, David; Zaias, Julia; Altman, Norman H

2008-11-01

Fenbendazole (FBZ) is an anthelmintic drug widely used to treat and prevent pinworm outbreaks in laboratory rodents. Although data in nonrodent species indicate possible effects of fenbendazole on the bone marrow and lymphocyte proliferation and function, little has been reported regarding possible effects on the rodent immune system. The purpose of the current study was to determine the effects of a therapeutic regimen of FBZ on immune parameters in BALB/c mice. Both 9-wk on-off and 5-wk continuous medicated feed protocols were assessed. No significant differences between normal and FBZ diet treated mice were observed in the following parameters: complete blood count, blood chemistry, quantitation of major T and B cell markers in spleen, quantitation of T cell markers in the thymus, spleen cell proliferation to T and B cell mitogens, bone marrow colony-forming cell assays, skin graft rejection, and primary and secondary humoral immune responses. These data indicate that FBZ treatment does not affect many standard broad measures of immune function.

The Modern Primitives: Applying New Technological Approaches to Explore the Biology of the Earliest Red Blood Cells

PubMed Central

Fraser, Stuart T.

2013-01-01

One of the most critical stages in mammalian embryogenesis is the independent production of the embryo's own circulating, functional red blood cells. Correspondingly, erythrocytes are the first cell type to become functionally mature during embryogenesis. Failure to achieve this invariably leads to in utero lethality. The recent application of technologies such as transcriptome analysis, flow cytometry, mutant embryo analysis, and transgenic fluorescent gene expression reporter systems has shed new light on the distinct erythroid lineages that arise early in development. Here, I will describe the similarities and differences between the distinct erythroid populations that must form for the embryo to survive. While much of the focus of this review will be the poorly understood primitive erythroid lineage, a discussion of other erythroid and hematopoietic lineages, as well as the cell types making up the different niches that give rise to these lineages, is essential for presenting an appropriate developmental context of these cells. PMID:24222861

Gap junction-mediated calcium waves define communication networks among murine postnatal neural progenitor cells.

PubMed

Lacar, Benjamin; Young, Stephanie Z; Platel, Jean-Claude; Bordey, Angélique

2011-12-01

In the postnatal neurogenic niche, two populations of astrocyte-like cells (B cells) persist, one acting as neural progenitor cells (NPCs, B1 cells) and one forming a structural boundary between the neurogenic niche and the striatum (B2 cells, niche astrocytes). Despite being viewed as two distinct entities, we found that B1 and B2 cells express the gap junction protein connexin 43 and display functional coupling involving 50-60 cells. Using neonatal electroporation to label slowly cycling radial glia-derived B1 cells, which send a basal process onto blood vessels, we further confirmed dye coupling between NPCs. To assess the functionality of the coupling, we used calcium imaging in a preparation preserving the three-dimensional architecture of the subventricular zone. Intercellular calcium waves were observed among B cells. These waves travelled bidirectionally between B1 and B2 cells and propagated on blood vessels. Inter-B-cell calcium waves were absent in the presence of a gap junction blocker but persisted with purinergic receptor blockers. These findings show that privileged microdomains of communication networks exist among NPCs and niche astrocytes. Such functional coupling between these two cell types suggests that niche astrocytes do not merely have a structural role, but may play an active role in shaping the behavior of NPCs. © 2011 The Authors. European Journal of Neuroscience © 2011 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.

Structure and function of the digestive system of solen grandis dunker

NASA Astrophysics Data System (ADS)

Sheng, Xiuzhen; Zhan, Wenbin; Ren, Sulian

2003-10-01

Structure and function of the digestive system of a bivalve mollusc, Solen grandis, were studied using light microscopy and histochemical methods. The wall of digestive tube consists of four layers: the mucosal epithelium, connective tissue, muscular and fibrosa or serosa (only in the portion of rectum) from the inner to the outer. The ciliated columnar epithelial cells, dispersed by cup-shaped mucous cells, rest on a thin base membrane. There are abundant blood spaces in connective tissue layer. The digestive diverticula are composed of multi-branched duct and digestive tubules. The digestive tubules are lined with digestive and basophilic secretory cells, and surrounded by a layer of smooth muscle fibers and connective tissues. Activities of acid and alkaline phosphatases, esterase and lipase are detected in the digestive cells, and the epithelia of stomach and intestine, suggesting that these cells are capable of intracellular digesting of food materials and absorbing. Besides, acid phosphatase and esterase activities are present in the posterior portion of esophagus. Phagocytes are abundant in blood spaces and the lumens of stomach and intestine, containing brown granules derived from the engulfed food materials. The present work indicates that phagocytes play important roles in ingestion and digestion of food materials, which is supported as well by the activities of acid phosphatase, esterase and lipase detected in blood spaces.

Sickle cell disease biochip: a functional red blood cell adhesion assay for monitoring sickle cell disease

PubMed Central

ALAPAN, YUNUS; KIM, CEONNE; ADHIKARI, ANIMA; GRAY, KAYLA E.; GURKAN-CAVUSOGLU, EVREN; LITTLE, JANE A.; GURKAN, UMUT A.

2016-01-01

Sickle cell disease (SCD) afflicts millions of people worldwide and is associated with considerable morbidity and mortality. Chronic and acute vaso-occlusion are the clinical hallmarks of SCD and can result in pain crisis, widespread organ damage, and early movtality. Even though the molecular underpinnings of SCD were identified more than 60 years ago, there are no molecular or biophysical markers of disease severity that are feasibly measured in the clinic. Abnormal cellular adhesion to vascular endothelium is at the root of vaso-occlusion. However, cellular adhesion is not currently evaluated clinically. Here, we present a clinically applicable microfluidic device (SCD biochip) that allows serial quantitative evaluation of red blood cell (RBC) adhesion to endothelium-associated protein-immobilized microchannels, in a closed and preprocessing-free system. With the SCD biochip, we have analyzed blood samples from more than 100 subjects and have shown associations between the measured RBC adhesion to endothelium-associated proteins (fibronectin and laminin) and individual RBC characteristics, including hemoglobin content, fetal hemoglobin concentration, plasma lactate dehydrogenase level, and reticulocyte count. The SCD biochip is a functional adhesion assay, reflecting quantitative evaluation of RBC adhesion, which could be used at baseline, during crises, relative to various long-term complications, and before and after therapeutic interventions. PMID:27063958

Numerical-experimental observation of shape bistability of red blood cells flowing in a microchannel

NASA Astrophysics Data System (ADS)

Guckenberger, Achim; Kihm, Alexander; John, Thomas; Wagner, Christian; Gekle, Stephan

Red blood cells flowing through capillaries assume a wide variety of different shapes owing to their high deformability. Predicting the realized shapes is a complex field as they are determined by the intricate interplay between the flow conditions and the membrane mechanics. In this work we construct the shape phase diagram of a single red blood cell with a physiological viscosity ratio flowing in a microchannel. We use both experimental in-vitro measurements as well as 3D numerical simulations to complement the respective other one. Numerically, we have easy control over the initial starting configuration and natural access to the full 3D shape. With this information we obtain the phase diagram as a function of initial position, starting shape and cell velocity. Experimentally, we measure the occurrence frequency of the different shapes as a function of the cell velocity to construct the experimental diagram which is in good agreement with the numerical observations. Two different major shapes are found, namely croissants and slippers. Notably, both shapes show coexistence at low (<1 mm/s) and high velocities (>3 mm/s) while in-between only croissants are stable. This pronounced bistability indicates that RBC shapes are not only determined by system parameters such as flow velocity or channel size, but also strongly depend on the initial conditions.

Dynamics and functions of CD4⁺CD25 (high) regulatory T lymphocytes in Chinese rhesus macaques during the early stage of infection with SIVmac239.

PubMed

Li, Shao-You; Xia, Hou-Jun; Dai, Zheng-Xi; Zhang, Gao-Hong; Fan, Bo; Li, Ming-Hua; Wang, Rui-Rui; Zheng, Yong-Tang

2012-05-01

CD4(+)CD25(high) regulatory T cells (Treg), which are a specialized subset of T cells, play an important role in the prevention of autoimmune diseases, maintenance of immune system homeostasis and tolerance to self-antigens. Chinese rhesus macaques (CRMs) are widely used in preclinical research on potential therapeutic drugs, vaccines and mechanisms of human diseases. However, the basic immunological characterization of Treg cells of CRMs has not been well established. To characterize Treg cells, peripheral blood of 43 adult CRMs was analyzed for CD4+ T lymphocytes by flow cytometry. It was found that Treg cells ranged from 1.52% to 11.1% of CD4+ T cells, and the average value was 5.7%. With our SIV-infected CRM model, through further studies, it was found that Treg cells in peripheral blood increased both in relative and absolute quantities. Moreover, Treg cells maintained their functions by suppressing Th1 cytokine secretion of their target cells. The results show that Treg cells might render cellular immunity against SIV viruses dysfunctional during the early stage after infection.

A comparison of cryopreservation methods: Slow-cooling vs. rapid-cooling based on cell viability, oxidative stress, apoptosis, and CD34+ enumeration of human umbilical cord blood mononucleated cells

PubMed Central

2011-01-01

Background The finding of human umbilical cord blood as one of the most likely sources of hematopoietic stem cells offers a less invasive alternative for the need of hematopoietic stem cell transplantation. Due to the once-in-a-life time chance of collecting it, an optimum cryopreservation method that can preserve the life and function of the cells contained is critically needed. Methods Until now, slow-cooling has been the routine method of cryopreservation; however, rapid-cooling offers a simple, efficient, and harmless method for preserving the life and function of the desired cells. Therefore, this study was conducted to compare the effectiveness of slow- and rapid-cooling to preserve umbilical cord blood of mononucleated cells suspected of containing hematopoietic stem cells. The parameters used in this study were differences in cell viability, malondialdehyde content, and apoptosis level. The identification of hematopoietic stem cells themselves was carried out by enumerating CD34+ in a flow cytometer. Results Our results showed that mononucleated cell viability after rapid-cooling (91.9%) was significantly higher than that after slow-cooling (75.5%), with a p value = 0.003. Interestingly, the malondialdehyde level in the mononucleated cell population after rapid-cooling (56.45 μM) was also significantly higher than that after slow-cooling (33.25 μM), with a p value < 0.001. The apoptosis level in rapid-cooling population (5.18%) was not significantly different from that of the mononucleated cell population that underwent slow-cooling (3.81%), with a p value = 0.138. However, CD34+ enumeration was much higher in the population that underwent slow-cooling (23.32 cell/μl) than in the one that underwent rapid-cooling (2.47 cell/μl), with a p value = 0.001. Conclusions Rapid-cooling is a potential cryopreservation method to be used to preserve the umbilical cord blood of mononucleated cells, although further optimization of the number of CD34+ cells after rapid-cooling is critically needed. PMID:21943045

Cellular Mechanisms of Somatic Stem Cell Aging

PubMed Central

Jung, Yunjoon

2014-01-01

Tissue homeostasis and regenerative capacity rely on rare populations of somatic stem cells endowed with the potential to self-renew and differentiate. During aging, many tissues show a decline in regenerative potential coupled with a loss of stem cell function. Cells including somatic stem cells have evolved a series of checks and balances to sense and repair cellular damage to maximize tissue function. However, during aging the mechanisms that protect normal cell function begin to fail. In this review, we will discuss how common cellular mechanisms that maintain tissue fidelity and organismal lifespan impact somatic stem cell function. We will highlight context-dependent changes and commonalities that define aging, by focusing on three age-sensitive stem cell compartments: blood, neural, and muscle. Understanding the interaction between extrinsic regulators and intrinsic effectors that operate within different stem cell compartments is likely to have important implications for identifying strategies to improve health span and treat age-related degenerative diseases. PMID:24439814

Transverse water relaxation in whole blood and erythrocytes at 3T, 7T, 9.4T, 11.7T and 16.4T; determination of intracellular hemoglobin and extracellular albumin relaxivities.

PubMed

Grgac, Ksenija; Li, Wenbo; Huang, Alan; Qin, Qin; van Zijl, Peter C M

2017-05-01

Blood is a physiological substance with multiple water compartments, which contain water-binding proteins such as hemoglobin in erythrocytes and albumin in plasma. Knowing the water transverse (R 2 ) relaxation rates from these different blood compartments is a prerequisite for quantifying the blood oxygenation level-dependent (BOLD) effect. Here, we report the Carr-Purcell-Meiboom-Gill (CPMG) based transverse (R 2CPMG ) relaxation rates of water in bovine blood samples circulated in a perfusion system at physiological temperature in order to mimic blood perfusion in humans. R 2CPMG values of blood plasma, lysed packed erythrocytes, lysed plasma/erythrocyte mixtures, and whole blood at 3 T, 7 T, 9.4 T, 11.7 T and 16.4 T were measured as a function of hematocrit or hemoglobin concentration, oxygenation, and CPMG inter-echo spacing (τ cp ). R 2CPMG in lysed cells showed a small τ cp dependence, attributed to the water exchange rate between free and hemoglobin-bound water to be much faster than τ cp . This was contrary to the tangential dependence in whole blood, where a much slower exchange between cells and blood plasma applies. Whole blood data were fitted as a function of τ cp using a general tangential correlation time model applicable for exchange as well as diffusion contributions to R 2CPMG , and the intercept R 20blood at infinitely short τ cp was determined. The R 20blood values at different hematocrit and the R 2CPMG values of lysed erythrocyte/plasma mixtures at different hemoglobin concentration were used to determine the relaxivity of hemoglobin inside the erythrocyte (r 2Hb ) and albumin (r 2Alb ) in plasma. The r 2Hb values obtained from lysed erythrocytes and whole blood were comparable at full oxygenation. However, while r 2Hb determined from lysed cells showed a linear dependence on oxygenation, this dependence became quadratic in whole blood. This possibly suggests an additional relaxation effect inside intact cells, perhaps due to hemoglobin proximity to the erythrocyte membrane. However, we cannot exclude that this is a consequence of the simple tangential model used to remove relaxation contributions from exchange and diffusion. The extensive data set presented should be useful for future theory development for the transverse relaxation of blood. Copyright © 2016 Elsevier Inc. All rights reserved.

Premature cell senescence and T cell receptor-independent activation of CD8T cells in Juvenile Idiopathic Arthritis*

PubMed Central

Dvergsten, Jeffrey A.; Mueller, Robert G.; Griffin, Patricia; Abedin, Sameem; Pishko, Allyson; Michel, Joshua J.; Rosenkranz, Margalit E.; Reed, Ann M.; Kietz, Daniel A.; Vallejo, Abbe N.

2013-01-01

Objectives CD8T cells lacking CD28 were originally reported by Wedderburn and colleagues as a characteristic feature of JIA, but the relevance of these unusual cells to JIA remains to be elucidated. Because of recent evidence that CD28 loss is typical of terminally differentiated lymphocytes, we examined for functional subsets of CD8T cells in JIA. Methods Following informed consent/assent, blood and/or waste synovial fluid were collected from children with definite diagnosis of JIA (n = 98). De-identified blood (n = 33) and cord blood (n = 13) samples from healthy donors were also collected. CD8T and CD4T cells were screened for novel receptors, and where indicated, bioassays were performed to determine functional relevance of the identified receptor. Results Patients had a naïve T cell compartment with shortened telomeres, and their entire T cell pool had reduced proliferative capacity. They had an over abundance of CD31+CD28null CD8T cells, which was a significant feature of oligoarticular JIA (n = 62) compared to polyarticular JIA (n = 36). CD31+CD28null CD8T cells had limited mitotic capacity, and expressed high levels of the senescence antigens γH2Ax and/or p16. Ligation of CD31, independent of the TCR, sufficiently induced tyrosine phosphorylation, vesicle exocytosis, and production of IFN-γ and IL-10. Conclusion These data provide the first evidence for cell senescence, represented by CD31+CD28null CD8T cells, in the pathophysiology of JIA. Activation of these unusual cells in a TCR-independent manner suggests they are maladaptive, and could be potential targets for immunotherapy. PMID:23686519

Inhibition of Aldehyde Dehydrogenase-Activity Expands Multipotent Myeloid Progenitor Cells with Vascular Regenerative Function.

PubMed

Cooper, Tyler T; Sherman, Stephen E; Kuljanin, Miljan; Bell, Gillian I; Lajoie, Gilles A; Hess, David A

2018-05-01

Blood-derived progenitor cell transplantation holds potential for the treatment of severe vascular diseases. Human umbilical cord blood (UCB)-derived hematopoietic progenitor cells purified using high aldehyde dehydrogenase (ALDH hi ) activity demonstrate pro-angiogenic functions following intramuscular (i.m.) transplantation into immunodeficient mice with hind-limb ischemia. Unfortunately, UCB ALDH hi cells are rare and prolonged ex vivo expansion leads to loss of high ALDH-activity and diminished vascular regenerative function. ALDH-activity generates retinoic acid, a potent driver of hematopoietic differentiation, creating a paradoxical challenge to expand UCB ALDH hi cells while limiting differentiation and retaining pro-angiogenic functions. We investigated whether inhibition of ALDH-activity during ex vivo expansion of UCB ALDH hi cells would prevent differentiation and expand progeny that retained pro-angiogenic functions after transplantation into non-obese diabetic/severe combined immunodeficient mice with femoral artery ligation-induced unilateral hind-limb ischemia. Human UCB ALDH hi cells were cultured under serum-free conditions for 9 days, with or without the reversible ALDH-inhibitor, diethylaminobenzaldehyde (DEAB). Although total cell numbers were increased >70-fold, the frequency of cells that retained ALDH hi /CD34+ phenotype was significantly diminished under basal conditions. In contrast, DEAB-inhibition increased total ALDH hi /CD34+ cell number by ≥10-fold, reduced differentiation marker (CD38) expression, and enhanced the retention of multipotent colony-forming cells in vitro. Proteomic analysis revealed that DEAB-treated cells upregulated anti-apoptotic protein expression and diminished production of proteins implicated with megakaryocyte differentiation. The i.m. transplantation of DEAB-treated cells into mice with hind-limb ischemia stimulated endothelial cell proliferation and augmented recovery of hind-limb perfusion. DEAB-inhibition of ALDH-activity delayed hematopoietic differentiation and expanded multipotent myeloid cells that accelerated vascular regeneration following i.m. transplantation in vivo. Stem Cells 2018;36:723-736. © AlphaMed Press 2018.

Endothelial Progenitor Cells Bind and Inhibit Platelet Function and Thrombus Formation

PubMed Central

Abou-Saleh, Haissam; Yacoub, Daniel; Théorêt, Jean-François; Gillis, Marc-Antoine; Neagoe, Paul-Eduard; Labarthe, Benoit; Théroux, Pierre; Sirois, Martin G.; Tabrizian, Maryam; Thorin, Eric; Merhi, Yahye

2013-01-01

Background Interactions of endothelial progenitor cells (EPCs) with vascular and blood cells contribute to vascular homeostasis. Although platelets promote the homing of EPCs to sites of vascular injury and their differentiation into endothelial cells, the functional consequences of such interactions on platelets remain unknown. Herein, we addressed the interactions between EPCs and platelets and their impact on platelet function and thrombus formation. Methods and Results Cultured on fibronectin in conditioned media, human peripheral blood mononuclear cells differentiated, within 10 days of culture, into EPCs, which uptake acetylated low-density lipoprotein, bind ulex-lectin, lack monocyte/leukocyte markers (CD14, P-selectin glycoprotein ligand-1, L-selectin), express progenitor/endothelial markers (CD34, vascular endothelial growth factor receptor-2, von Willebrand factor, and vascular endothelial cadherin), and proliferate in culture. These EPCs bound activated platelets via CD62P and inhibited its translocation, glycoprotein IIb/IIIa activation, aggregation, and adhesion to collagen, mainly via prostacyclin secretion. Indeed, this was associated with upregulation of cyclooxygenase-2 and inducible nitric oxide synthase. However, the effects on platelets in vitro were reversed by cyclooxygenase and cyclooxygenase-2 inhibition but not by nitric oxide or inducible nitric oxide synthase inhibition. Moreover, in a ferric chloride–induced murine arterial thrombosis model, injection of EPCs led to their incorporation into sites of injury and impaired thrombus formation, leading to an incomplete occlusion with 50% residual flow. Conclusions Peripheral blood mononuclear cell– derived EPCs bind platelets via CD62P and inhibit platelet activation, aggregation, adhesion to collagen, and thrombus formation, predominantly via upregulation of cyclooxygenase-2 and secretion of prostacyclin. These findings add new insights into the biology of EPCs and define their potential roles in regulating platelet function and thrombosis. PMID:19917882

Age-Associated Decline in Dendritic Cell Function and the Impact of Mediterranean Diet Intervention in Elderly Subjects.

PubMed

Clements, Sarah J; Maijo, Monica; Ivory, Kamal; Nicoletti, Claudio; Carding, Simon R

2017-01-01

Aging is accompanied by increased susceptibility to infection and age-associated chronic diseases. It is also associated with reduced vaccine responses, which is often attributed to immunosenescence and the functional decline of the immune system. Immunosenescence is characterized by a chronic, low-grade, inflammatory state termed inflammaging. Habitants of Mediterranean (MED) regions maintain good health into old age; often attributed to MED diets. Adoption of a MED-diet by elderly subjects, in Norfolk (UK), may improve immune responses of these individuals and in particular, dendritic cell (DC) function. A total of 120 elderly subjects (65-79 years old) recruited onto the Nu-AGE study, a multicenter European dietary study specifically addressing the needs of the elderly, across five countries, and were randomized to the control or MED-diet groups, for one year. Blood samples were taken pre- and post-intervention for DC analysis and were compared with each other, and to samples obtained from 45 young (18-40 years old) subjects. MED-diet compliance was assessed using high performance liquid chromatography-with tandem mass spectrometry analysis of urine samples. Immune cell and DC subset numbers and concentrations of secreted proteins were determined by flow cytometric analysis. As expected, reduced myeloid DC numbers were observed in blood samples from elderly subjects compared with young. The elevated secretion of the adipokine, resistin, after ex vivo stimulation of peripheral blood mononuclear cells from elderly subjects, was significantly reduced after MED-diet intervention. This study provides further evidence of numerical and functional effects of aging on DCs. The MED-diet showed potential to impact on the aging immune cells investigated and could provide an economical approach to address problems associated with our aging population.

Regulatory T Cells in Peripheral Blood and Cerebrospinal Fluid of Syphilis Patients with and without Neurological Involvement

PubMed Central

Li, Kang; Wang, Cuini; Lu, Haikong; Gu, Xin; Guan, Zhifang; Zhou, Pingyu

2013-01-01

Background Syphilis, a sexually transmitted disease caused by spirochetal bacterium Treponema pallidum, can progress to affect the central nervous system, causing neurosyphilis. Accumulating evidence suggest that regulatory T cells (Tregs) may play an important role in the pathogenesis of syphilis. However, little is known about Treg response in neurosyphilis. Methodology/Principal Findings We analyzed Treg frequencies and Transforming Growth Factor-β (TGF-β) levels in the blood and CSF of 431 syphilis patients without neurological involvement, 100 neurosyphilis patients and 100 healthy donors. Suppressive function of Tregs in peripheral blood was also assessed. Among syphilis patients without neurological involvement, we found that secondary and serofast patients had increased Treg percentages, suppressive function and TGF-β levels in peripheral blood compared to healthy donors. Serum Rapid Plasma Reagin (RPR) titers were positively correlated with Treg numbers in these patients. Compared to these syphilis patients without neurological involvement, neurosyphilis patients had higher Treg frequency in peripheral blood. In the central nervous system, neurosyphilis patients had higher numbers of leukocytes in CSF compared to syphilis patients without neurological involvement. CD4+ T cells were the predominant cell type in the inflammatory infiltrates in CSF of neurosyphilis patients. Interestingly, among these neurosyphilis patients, a significant decrease in CSF CD4+ CD25high Treg percentage and number was observed in symptomatic neurosyphilis patients compared to those of asymptomatic neurosyphilis patients, which may be associated with low CSF TGF-β levels. Conclusions Our findings suggest that Tregs might play an important role in both bacterial persistence and neurologic compromise in the pathogenesis of syphilis. PMID:24244772

Normal platelet function in platelet concentrates requires non-platelet cells: a comparative in vitro evaluation of leucocyte-rich (type 1a) and leucocyte-poor (type 3b) platelet concentrates

PubMed Central

Parrish, William R; Roides, Breana; Hwang, Julia; Mafilios, Michael; Story, Brooks; Bhattacharyya, Samir

2016-01-01

Background Therapeutic success of platelet-rich plasma (PRP) may vary based on the composition and preparation method. The objective of this study was to evaluate the cellular components of platelet concentrates produced by a leucocyte-rich (LR-PRP) and a leucocyte-poor PRP systems (LP-PRP). Methods Parameters evaluated included platelet recovery, platelet concentration, red blood cell (RBC) and white blood cell (WBC) composition, platelet growth factor release and stimulation of human tendon cell proliferation in vitro. Results Platelet recoveries were 52% for LP-PRP and 89% for LR-PRP. LR-PRP demonstrated greater reproducibility with a 4.2% coefficient of variation (CV) compared with 19.4% for LP-PRP (p<0.001). LR-PRP demonstrated a greater increase in platelet concentration (7.9-fold) than LP-PRP (2.2-fold; p<0.001). LP-PRP showed 5.0-fold reductions in WBCs, while LR-PRP showed a 4.0-fold increase (p<0.001). LP-PRP reduced RBCs to a haematocrit of 0.25, while LR-PRP reduced haematocrit to 11.8. LP-PRP did not coagulate robustly on reactivation with CaCl2, and released significantly lower levels of epidermal growth factor (EGF) and transforming growth factor β1 (TGF-β1) than whole blood (p<0.03). LP-PRP also did not stimulate tendon cell proliferation greater than whole blood. In contrast, LR-PRP showed increases in each growth factor on activation with CaCl2 (p<0.01) and stimulated greater proliferation (p<0.05) compared with whole blood. Forced activation of LP-PRP with exogenous thrombin rescued the coagulation deficiency and induced greater growth factor release than comparable whole blood (p<0.03). Conclusions These data suggest that non-platelet cellular components in platelet concentrates are important for proper platelet function, including thrombin generation, growth factor release and clot retraction. PMID:27900155

Normal platelet function in platelet concentrates requires non-platelet cells: a comparative in vitro evaluation of leucocyte-rich (type 1a) and leucocyte-poor (type 3b) platelet concentrates.

PubMed

Parrish, William R; Roides, Breana; Hwang, Julia; Mafilios, Michael; Story, Brooks; Bhattacharyya, Samir

2016-01-01

Therapeutic success of platelet-rich plasma (PRP) may vary based on the composition and preparation method. The objective of this study was to evaluate the cellular components of platelet concentrates produced by a leucocyte-rich (LR-PRP) and a leucocyte-poor PRP systems (LP-PRP). Parameters evaluated included platelet recovery, platelet concentration, red blood cell (RBC) and white blood cell (WBC) composition, platelet growth factor release and stimulation of human tendon cell proliferation in vitro. Platelet recoveries were 52% for LP-PRP and 89% for LR-PRP. LR-PRP demonstrated greater reproducibility with a 4.2% coefficient of variation (CV) compared with 19.4% for LP-PRP (p<0.001). LR-PRP demonstrated a greater increase in platelet concentration (7.9-fold) than LP-PRP (2.2-fold; p<0.001). LP-PRP showed 5.0-fold reductions in WBCs, while LR-PRP showed a 4.0-fold increase (p<0.001). LP-PRP reduced RBCs to a haematocrit of 0.25, while LR-PRP reduced haematocrit to 11.8. LP-PRP did not coagulate robustly on reactivation with CaCl 2 , and released significantly lower levels of epidermal growth factor (EGF) and transforming growth factor β1 (TGF-β1) than whole blood (p<0.03). LP-PRP also did not stimulate tendon cell proliferation greater than whole blood. In contrast, LR-PRP showed increases in each growth factor on activation with CaCl 2 (p<0.01) and stimulated greater proliferation (p<0.05) compared with whole blood. Forced activation of LP-PRP with exogenous thrombin rescued the coagulation deficiency and induced greater growth factor release than comparable whole blood (p<0.03). These data suggest that non-platelet cellular components in platelet concentrates are important for proper platelet function, including thrombin generation, growth factor release and clot retraction.

Mesenchymal stem cells inhibit dendritic cell differentiation and function by preventing entry into the cell cycle.

PubMed

Ramasamy, Rajesh; Fazekasova, Henrietta; Lam, Eric W-F; Soeiro, Inês; Lombardi, Giovanna; Dazzi, Francesco

2007-01-15

Mesenchymal stem cells (MSCs) play a crucial role in hematopoietic development and have been shown to exert a powerful immunosuppressive effect. In this study, we investigated the effect of bone marrow MSC on the differentiation and function of peripheral blood monocytes into dendritic cells (DCs). Human MSCs, generated from normal bone marrow, were added to peripheral blood monocytes stimulated in vitro with granulocyte-macrophage colony stimulating factor and interleukin-4 to become DCs. Monocytes were then examined for the expression of markers characteristic of DCs and their ability to stimulate allogeneic T cells. In addition, the effect of MSCs on the cell cycle of monocyte-derived DCs and the expression of various cell cycle proteins were analyzed by cytometric analysis and Western blotting with specific antibodies. MSCs blocked the differentiation of monocytes into DCs and impaired their antigen-presenting ability. This resulted from a block of monocytes from entering the G1 phase of the cell cycle with a progressive number of cells accumulating in the G0 phase. Cyclin D2 was downregulated. However, differently from what was observed in T-cells stimulated in the presence of MSCs, the expression of p27 was found decreased, suggesting the involvement of similar but not identical pathways. We conclude that MSCs impair monocyte differentiation and function by interfering with the cell cycle. These findings imply that MSC-induced immunosuppression might be a side product of a more general antiproliferative effect.

Butyrate protects against disruption of the blood-milk barrier and moderates inflammatory responses in a model of mastitis induced by lipopolysaccharide.

PubMed

Wang, Jing-Jing; Wei, Zheng-Kai; Zhang, Xu; Wang, Ya-Nan; Fu, Yun-He; Yang, Zheng-Tao

2017-11-01

Short-chain fatty acids are fermentation end products produced by gut bacteria, which have been shown to ameliorate inflammatory bowel diseases and allergic asthma. However, the mechanism involved remains largely unknown. Here, we investigate the protective effects and mechanisms of sodium butyrate (SB) on LPS-induced mastitis model. Effects of increasing doses of SB on blood-milk barrier function and inflammation are studied in BALB/c mice with LPS-induced mastitis. The underlying mechanisms of anti-inflammatory effects of SB were further investigated in LPS-stimulated mouse mammary epithelial cells (mMECs). The results show that SB decreased LPS-induced disruption in mammary tissues, infiltration of inflammatory cells and the levels of TNF-α, IL-6 and IL-1β. SB up-regulated the tight junction proteins occludin and claudin-3 and reduced blood-milk barrier permeability in LPS-induced mastitis. Studies in vitro revealed that SB inhibited LPS-induced inflammatory response by inhibition of the NF-κB signalling pathway and histone deacetylases in LPS-stimulated mMECs. In our model, SB protected against LPS-induced mastitis by preserving blood-milk barrier function and depressing pro-inflammatory responses, suggesting the potential use of SB as a prophylactic agent to protect blood-milk barrier function in mastitis. © 2017 The British Pharmacological Society.

In vitro transdifferentiation of human peripheral blood mononuclear cells to photoreceptor-like cells

PubMed Central

Komuta, Yukari; Ishii, Toshiyuki; Kaneda, Makoto; Ueda, Yasuji; Miyamoto, Kiyoko; Toyoda, Masashi; Umezawa, Akihiro

2016-01-01

ABSTRACT Direct reprogramming is a promising, simple and low-cost approach to generate target cells from somatic cells without using induced pluripotent stem cells. Recently, peripheral blood mononuclear cells (PBMCs) have attracted considerable attention as a somatic cell source for reprogramming. As a cell source, PBMCs have an advantage over dermal fibroblasts with respect to the ease of collecting tissues. Based on our studies involving generation of photosensitive photoreceptor cells from human iris cells and human dermal fibroblasts by transduction of photoreceptor-related transcription factors via retrovirus vectors, we transduced these transcription factors into PBMCs via Sendai virus vectors. We found that retinal disease-related genes were efficiently detected in CRX-transduced cells, most of which are crucial to photoreceptor functions. In functional studies, a light-induced inward current was detected in some CRX-transduced cells. Moreover, by modification of the culture conditions including additional transduction of RAX1 and NEUROD1, we found a greater variety of retinal disease-related genes than that observed in CRX-transduced PBMCs. These data suggest that CRX acts as a master control gene for reprogramming PBMCs into photoreceptor-like cells and that our induced photoreceptor-like cells might contribute to individualized drug screening and disease modeling of inherited retinal degeneration. PMID:27170256

Synthetic Capillaries to Control Microscopic Blood Flow

NASA Astrophysics Data System (ADS)

Sarveswaran, K.; Kurz, V.; Dong, Z.; Tanaka, T.; Penny, S.; Timp, G.

2016-02-01

Capillaries pervade human physiology. The mean intercapillary distance is only about 100 μm in human tissue, which indicates the extent of nutrient diffusion. In engineered tissue the lack of capillaries, along with the associated perfusion, is problematic because it leads to hypoxic stress and necrosis. However, a capillary is not easy to engineer due to its complex cytoarchitecture. Here, it is shown that it is possible to create in vitro, in about 30 min, a tubular microenvironment with an elastic modulus and porosity consistent with human tissue that functionally mimicks a bona fide capillary using “live cell lithography”(LCL) to control the type and position of cells on a composite hydrogel scaffold. Furthermore, it is established that these constructs support the forces associated with blood flow, and produce nutrient gradients similar to those measured in vivo. With LCL, capillaries can be constructed with single cell precision—no other method for tissue engineering offers such precision. Since the time required for assembly scales with the number of cells, this method is likely to be adapted first to create minimal functional units of human tissue that constitute organs, consisting of a heterogeneous population of 100-1000 cells, organized hierarchically to express a predictable function.

Synthetic Capillaries to Control Microscopic Blood Flow.

PubMed

Sarveswaran, K; Kurz, V; Dong, Z; Tanaka, T; Penny, S; Timp, G

2016-02-24

Capillaries pervade human physiology. The mean intercapillary distance is only about 100 μm in human tissue, which indicates the extent of nutrient diffusion. In engineered tissue the lack of capillaries, along with the associated perfusion, is problematic because it leads to hypoxic stress and necrosis. However, a capillary is not easy to engineer due to its complex cytoarchitecture. Here, it is shown that it is possible to create in vitro, in about 30 min, a tubular microenvironment with an elastic modulus and porosity consistent with human tissue that functionally mimicks a bona fide capillary using "live cell lithography"(LCL) to control the type and position of cells on a composite hydrogel scaffold. Furthermore, it is established that these constructs support the forces associated with blood flow, and produce nutrient gradients similar to those measured in vivo. With LCL, capillaries can be constructed with single cell precision-no other method for tissue engineering offers such precision. Since the time required for assembly scales with the number of cells, this method is likely to be adapted first to create minimal functional units of human tissue that constitute organs, consisting of a heterogeneous population of 100-1000 cells, organized hierarchically to express a predictable function.

Circulating tumor cells (CTCs) from metastatic breast cancer patients linked to decreased immune function and response to treatment.

PubMed

Green, Taryn L; Cruse, Julius M; Lewis, Robert E; Craft, Barbara S

2013-10-01

We aimed to examine the use of circulating tumor cells (CTCs) as an effective measure of treatment efficacy and immune system function in metastatic breast cancer patients. CTCs are believed to be indicators of residual disease and thus pose an increased risk of metastasis and poorer outcomes to those patients who are CTC-positive. We obtained peripheral blood samples from 45 patients previously diagnosed with metastatic disease originating in the breast. Using TLR agonists that bind TLR ligands and upregulate immune effects versus unstimulated cells, we calculated a percent specific lysis using chromium-51 assay to illustrate the functional abilities of patient natural killer (NK) cells. We found those with greater than 5 CTCs per 7.5 mL blood had significantly decreased responses by their immune cells when compared with those patients who had 5 CTCs or less. We furthermore found a correlation between disease progression and CTC-positive patients, indicating that those who have a positive test should be closely monitored by their clinician. CTCs represent an exciting new clinical opportunity that will ideally utilize their low invasiveness and quick turnaround time to best benefit clinical scenarios. © 2013.

T cell subsets in cord blood are influenced by maternal allergy and associated with atopic dermatitis.

PubMed

Fu, Yujing; Lou, Hongfei; Wang, Chengshuo; Lou, Wei; Wang, Yang; Zheng, Tao; Zhang, Luo

2013-03-01

This study aimed to investigate the influence of maternal allergy on cord blood regulatory and effector T cells and to evaluate their role as a predictor of atopic dermatitis (AD) during the first 2 yr of life. Seventy mother-infant pairs were recruited in this prospective birth cohort study (21 allergic and 49 non-allergic mothers). Cord blood samples were collected and assayed for the percentage of regulatory T cells (Treg), interferon-γ (IFN-γ), and interleukin-4 (IL-4) producing T cells (Th1 and Th2, respectively) using flow cytometry. Experiments were undertaken to assess the function of cord blood CD4(+) CD25(+) CD127(-) Treg cells by cell proliferation and cytokine responses. Their offspring at the age of 2 yr old were evaluated by dermatologists to determine whether they had AD. During the first 2 yr of life, 15.7% of the children developed a physician-diagnosed AD. A significantly increased percentage of Th2 cell was observed in cord blood of newborns with maternal allergy. Treg/Th2 ratio significantly decreased among the offspring of allergic mothers. Treg cell-associated suppression of Th2 response was attenuated in Der p1-stimulated CD4(+) CD25(-) T cells from the offspring of allergic mothers. Children with reduced Th1/Th2 (p = 0.001, OR = 0.37) and Treg/Th2 (p = 0.001, OR = 0.47) ratio in cord blood had a higher risk of developing AD. Maternal allergic status is associated with increased percentage of IL-4(+) CD4(+) T cells and a reduced Treg/Th2 ratio in cord blood at their children's birth, which may predispose to an increased risk for developing AD. © 2013 John Wiley & Sons A/S. Published by Blackwell Publishing Ltd.

Effect of human milk on blood and bone marrow cells in a malnourished mice model; comparative study with cow milk.

PubMed

García, Isabel; Salva, Susana; Zelaya, Hortensia; Villena, Julio; Agüero, Graciela

2013-11-01

It has been demonstrated that the alterations caused by nutrient deficiency can be reverted by adequate nutritional repletion. To perform comparative studies between human and cow milks in order to evaluate the impact of both milks on the recovery of blood and bone marrow cells affected in malnourished mice. Weaned mice were malnourished after consuming a protein free diet for 21 days. Malnourished mice received cow or human milk (CM or HM) for 7 or 14 consecutive days. During the period of administration of milk, the mice consumed the protein free diet ad libitum. The malnourished control (MNC) group received only protein free diet whereas the wellnourished control (WNC) mice consumed the balanced conventional diet. Both milks normalized serum albumin levels and improved thymus weight. Human milk was less effective than cow milk to increase body weight and serum transferrin levels. In contrast, human milk was more effective than cow milk to increase the number of leukocytes (WNC: 6.90 ± 1.60a; MNC: 2.80 ± 0.90b; CM 7d: 3.74 ± 1.10b; HM 7d: 7.16 ± 1.90a; CM 14d: 4.35 ± 1.20b; HM 14d: 6.75 ± 1.20a (109/L); p < 0.05) and lymphocytes (WNC: 5.80 ± 0.36a; MNC: 1.80 ± 0.40b; CM 7d: 2.50 ± 0.30b; HM 7d: 4.20 ± 0.50c; CM 14d: 3.30 ± 0.31d; HM 14d: 4.70 ± 0.28c (109/L); p < 0.05) in peripheral blood. Both milks induced an increment in mitotic pool cells in bone marrow and α-naphthyl butyrate esterase positive cells in peripheral blood. They also normalized phagocytic function in blood neutrophils and oxidative burst in peritoneal cells. Both milks were equally effective to exert favorable effects on the number of the bone marrow cells and the functions of the blood and peritoneal cells involved in immune response. However, only human milk normalized the number of leukocytes and increased the number of neutrophils in peripheral blood. Copyright AULA MEDICA EDICIONES 2013. Published by AULA MEDICA. All rights reserved.

Menstrual blood closely resembles the uterine immune micro-environment and is clearly distinct from peripheral blood.

PubMed

van der Molen, R G; Schutten, J H F; van Cranenbroek, B; ter Meer, M; Donckers, J; Scholten, R R; van der Heijden, O W H; Spaanderman, M E A; Joosten, I

2014-02-01

Is menstrual blood a suitable source of endometrial derived lymphocytes? Mononuclear cells isolated from menstrual samples (menstrual blood mononuclear cells (MMC)) are clearly distinct from peripheral blood mononuclear cells (PBMC) and show a strong resemblance with biopsy-derived endometrial mononuclear cells. A critical event in the onset of pregnancy is the implantation of the embryo in the uterine wall. The immune cell composition in the endometrium at the time of implantation is considered pivotal for success. Despite advancing knowledge on the composition of the immune cell population in the uterus, the role of endometrial immune cells in reproductive disorders is still not fully resolved, mainly due to the fact that this type of research requires invasive techniques. Here, we collected menstrual fluid and validated this unique non-invasive technique to obtain and study the endometrium-derived immune cells which would be present around the time of implantation. Five healthy non-pregnant females with regular menstruation cycles and not using oral contraceptives collected their menstrual blood using a menstrual cup in five consecutive cycles. Sampling took place over the first 3 days of menses, with 12 h intervals. Peripheral blood samples, taken before and after each menstruation, were obtained for comparative analysis. MMC and PBMC samples were characterized for the different lymphocyte subsets by flow cytometry, with emphasis on NK cells and T cells. Next, the functional capacity of the MMC-derived NK cells was determined by measuring intracellular production of IFN-γ, granzyme B and perforin after culture in the presence of IL-2 and IL-15. In support of their endometrial origin, MMC samples contained the typical composition of mononuclear cells expected of endometrial tissue, were phenotypically similar to the reported phenotype for biopsy-derived endometrial cells, and were distinct from PBMC. Increased percentages of NK cells and decreased percentages of T cells were found in MMC when compared with PBMC from the same female. The MMC-derived NK cells were pre-dominantly CD56(bright)/CD16(-), in contrast to the primarily CD56(dim)/CD16(+) peripheral blood NK cells. MMC-derived NK cells expressed CD103, indicating their mucosal origin. In addition, the pattern of natural cytotoxicity receptor (NCR) expression in MMC-derived NK cells was comparable with that in endometrial biopsy-derived NK cells. Compared with PBMC, the NKp30 expression was decreased, while the percentage of NKp44 positive cells was increased in MMC samples. CXCR3 and CXCR4 were hardly expressed by MMC-derived NK cells, indicating that these cells are not of PBMC origin. NK cells from MMC samples were functional as shown by their capacity to produce IFN-γ, granzyme B and perforin, upon stimulation with IL-2 and IL-15. MMC-derived T cells revealed an increased expression of CD103, CD69 and CXCR4 compared with PBMC-derived T cells. Importantly, MMC collection using a menstrual cup proved highly reliable and reproducible between women and between cycles. Based on the parameters we studied, MMC appear similar to biopsy-derived endometrial mononuclear cells. However, sampling is not done at the exact same time in the menstrual cycle, and thus we cannot exclude some, as yet undetected, differences. Also, it should be considered that for some women, the use of the menstrual cup may be unpleasant. Menstrual blood may be a source of endometrial cells and may create new opportunities to study uterine immunological cells in fertility issues. No external funding was obtained for the present study. None of the authors have any conflict of interest to declare. NA.

Endothelial nitric oxide synthase in red blood cells: Key to a new erythrocrine function?☆

PubMed Central

Cortese-Krott, Miriam M.; Kelm, Malte

2014-01-01

Red blood cells (RBC) have been considered almost exclusively as a transporter of metabolic gases and nutrients for the tissues. It is an accepted dogma that RBCs take up and inactivate endothelium-derived NO via rapid reaction with oxyhemoglobin to form methemoglobin and nitrate, thereby limiting NO available for vasodilatation. Yet it has also been shown that RBCs not only act as “NO sinks”, but exert an erythrocrine function – i.e an endocrine function of RBC – by synthesizing, transporting and releasing NO metabolic products and ATP, thereby potentially controlling systemic NO bioavailability and vascular tone. Recent work from our and others laboratory demonstrated that human RBCs carry an active type 3, endothelial NO synthase (eNOS), constitutively producing NO under normoxic conditions, the activity of which is compromised in patients with coronary artery disease. In this review we aim to discuss the potential role of red cell eNOS in RBC signaling and function, and to critically revise evidence to this date showing a role of non-endothelial circulating eNOS in cardiovascular pathophysiology. PMID:24494200

Expression of NK cell receptors on decidual T cells in human pregnancy.

PubMed

Tilburgs, Tamara; van der Mast, Barbara J; Nagtzaam, Nicole M A; Roelen, Dave L; Scherjon, Sicco A; Claas, Frans H J

2009-06-01

Specific receptors enable NK cells to discriminate between cells with normal expression of MHC class I and cells that have low or absent expression of MHC class I molecules. In addition to NK cells, these receptors can be expressed on T cell subsets, mainly on CD8+ T cells but also on gammadeltaTCR+ T cells and CD4+ T cells. Although the function of NK cell receptor expression on T cells is not completely understood, various studies have shown that they are involved in down regulation of T cell receptor (TCR)-mediated activation and influence effector functions, like cytotoxicity and cytokine production. The aim of this study was to analyze expression of NK cell receptors on peripheral blood and decidual T cells during human pregnancy using flow cytometry. We demonstrate that a proportion of decidual T cells express HLA-C specific killer immunoglobulin-like receptors (KIRs). Furthermore, a small proportion of decidual T cells express the HLA-E specific CD94-NKG2A inhibitory and CD94-NKG2C activating receptors. Decidual KIR+ and CD94-NKG2+ T cells mainly display a CD3+CD4-CD8- phenotype. However, decidual tissue also contains higher percentages of KIR and CD94-NKG2 expressing CD4+ and CD8+ T cells compared to peripheral blood. So far, the functional capacities of decidual T cells expressing the NK cell receptors are unknown but NK cell receptor expression on decidual T cells may provide an alternative means by which decidual T cells distinguish self (maternal) cells from allogeneic fetal cells, and act to modulate the decidual immune response.

Ultracytochemical visualization of calcium distribution in heart cells and erythrocytes of zebrafish Danio rerio.

PubMed

Niksirat, Hamid; Steinbach, Christoph

2018-05-24

Detection of patterns of subcellular calcium distribution in the cardiovascular system can contribute to understanding its role in cardiac and blood function. The present study localized calcium in heart atrium, ventricle, and bulbus arteriosus as well as in erythrocytes of zebrafish Danio rerio using an oxalate-pyroantimonate technique combined with transmission electron microscopy. Intracellular calcium stores were detected in caveolae, mitochondria, and the nuclei of several zebrafish cardiac cell types. Melanin pigmentation containing calcium stores was detected in the pericardial cavity. Melanin might be an extracellular source of calcium for heart beating and/or a lubricant to prevent friction during beating process. Calcium deposits were also detected in the plasma membrane, cytoplasm and nucleus of erythrocytes as well as in blood plasma. Possible exchange of calcium between erythrocytes and blood plasma was observed. Interactions of such calcium stores and possible contribution of extracellular calcium stores such as melanin pigmentation to supply calcium for vital functions of heart cells should be addressed in future studies. Copyright © 2018 Elsevier Ltd. All rights reserved.

Avian macrophage: effector functions in health and disease.

PubMed

Qureshi, M A; Heggen, C L; Hussain, I

2000-01-01

Monocytes-macrophages, cells belonging to the mononuclear phagocytic system, are considered as the first line of immunological defense. Being mobile scavenger cells, macrophages participate in innate immunity by serving as phagocytic cells. These cells arise in the bone marrow and subsequently enter the blood circulation as blood monocytes. Upon migration to various tissues, monocytes mature and differentiate into tissue macrophages. Macrophages then initiate the 'acquired' immune response in their capacity as antigen processing and presenting cells. While responding to their tissue microenvironment or exogenous antigenic challenge, macrophages may secrete several immunoregulatory cytokines or metabolites. Being the first line of immunological defense, macrophages therefore represent an important step during interaction with infectious agents. The outcome of the macrophage-pathogen interaction depends upon several factors including the stage of macrophage activation, the nature of the infectious agent, the level of genetic control on macrophage function as well as environmental and nutritional factors that may modulate macrophage activation and functions. Research in avian macrophages has lagged behind that in mammals. This has been largely due to the lack of harvestable resident macrophages from the chicken peritoneal cavity. However, the development of elicitation protocols to harvest inflammatory abdominal macrophages and the availability of transformed chicken macrophage cell lines, has enabled researchers to address several questions related to chicken macrophage biology and function in health and disease. In this manuscript the basic profiles of several macrophage effector functions are described. In addition, the interaction of macrophages with various pathogens as well as the effect of genetic and environmental factors on macrophage functional modulation is described.

Effect of lignin supplementation of a diet contaminated with Fusarium mycotoxins on blood and intestinal lymphocyte subpopulations in chickens.

PubMed

Revajová, Viera; Levkut, Mikuláš; Levkutová, Mária; Bořutová, Radka; Grešaková, Lubomíra; Košiková, Božena; Leng, Lubomír

2013-09-01

The objective of the study was to investigate the effects of lignin supplementation of a diet contaminated with the Fusarium mycotoxins deoxynivalenol (DON) and zearalenone (ZEA) on peripheral blood leukocytes and duodenal immunocompetent cells in broiler chickens. From day 1 after hatching, all chickens were fed an identical control diet for two weeks. Then chickens of Group 1 continued to be fed the control diet, whereas Group 2 was fed the same diet supplemented with lignin at 0.5% level. Simultaneously, Group 3 started to receive a diet contaminated with DON (2.95 mg kg-1) and ZEA (1.59 mg kg-1), while Group 4 received an identical contaminated diet supplemented with 0.5% lignin for further two weeks. Samples of blood and duodenal tissue were collected from 6 birds of each group at 4 weeks of age. Neither counts of white blood cells nor phagocytic function in the peripheral blood were significantly affected in the mycotoxin- and/or lignin-treated birds. As compared to the control, increased numbers of IgM-bearing cells were found in the peripheral blood in Group 3 fed the contaminated diet (P < 0.05) and in Group 4 given the contaminated diet supplemented with lignin (P < 0.01). While the contaminated diet led to reduced numbers of duodenal CD4+ cells, in Group 2 treated only with lignin the number of duodenal CD4+ cells was increased. Lignin enrichment of the contaminated diet did not eliminate the mycotoxin-induced reduction in the number of duodenal CD4+ cells. The results suggest that dietary supplementation of lignin as an indigestible compound to poultry feed may increase the density of some intestinal immunocompetent cells without exerting effects on that in the peripheral blood. However, when added to a diet contaminated with Fusarium mycotoxins, lignin did not prevent the mycotoxin-induced changes in the numbers of blood and intestinal immunocompetent cells.

Granulocytic myeloid-derived suppressor cells from human cord blood modulate T-helper cell response towards an anti-inflammatory phenotype.

PubMed

Köstlin, Natascha; Vogelmann, Margit; Spring, Bärbel; Schwarz, Julian; Feucht, Judith; Härtel, Christoph; Orlikowsky, Thorsten W; Poets, Christian F; Gille, Christian

2017-09-01

Infections are a leading cause of perinatal morbidity and mortality. The outstandingly high susceptibility to infections early in life is mainly attributable to the compromised state of the neonatal immune system. One important difference to the adult immune system is a bias towards T helper type 2 (Th2) responses in newborns. However, mechanisms regulating neonatal T-cell responses are incompletely understood. Granulocytic myeloid-derived suppressor cells (GR-MDSC) are myeloid cells with a granulocytic phenotype that suppress various functions of other immune cells and accumulate under physiological conditions during pregnancy in maternal and fetal blood. Although it has been hypothesized that GR-MDSC accumulation during fetal life could be important for the maintenance of maternal-fetal tolerance, the influence of GR-MDSC on the immunological phenotype of neonates is still unclear. Here, we investigated the impact of GR-MDSC isolated from cord blood (CB-MDSC) on the polarization of Th cells. We demonstrate that CB-MDSC inhibit Th1 responses and induced Th2 responses and regulatory T (Treg) cells. Th1 inhibition was cell-contact dependent and occurred independent of other cell types, while Th2 induction was mediated independently of cell contact through expression of ArgI and reactive oxygen species by CB-MDSC and partially needed the presence of monocytes. Treg cell induction by CB-MDSC also occurred cell-contact independently but was partially mediated through inducible nitric oxide synthase. These results point towards a role of MDSC in regulating neonatal immune responses. Targeting MDSC function in neonates could be a therapeutic opportunity to improve neonatal host defence. © 2017 John Wiley & Sons Ltd.

Transplant of Hepatocytes, Undifferentiated Mesenchymal Stem Cells, and In Vitro Hepatocyte-Differentiated Mesenchymal Stem Cells in a Chronic Liver Failure Experimental Model: A Comparative Study.

PubMed

El Baz, Hanan; Demerdash, Zeinab; Kamel, Manal; Atta, Shimaa; Salah, Faten; Hassan, Salwa; Hammam, Olfat; Khalil, Heba; Meshaal, Safa; Raafat, Inas

2018-02-01

Liver transplant is the cornerstone line of treatment for chronic liver diseases; however, the long list of complications and obstacles stand against this operation. Searching for new modalities for treatment of chronic liver illness is a must. In the present research, we aimed to compare the effects of transplant of undifferentiated human mesenchymal stem cells, in vitro differentiated mesenchymal stem cells, and adult hepatocytes in an experimental model of chronic liver failure. Undifferentiated human cord blood mesenchymal stem cells were isolated, pro-pagated, and characterized by morphology, gene expression analysis, and flow cytometry of surface markers and in vitro differentiated into hepatocyte-like cells. Rat hepatocytes were isolated by double perfusion technique. An animal model of chronic liver failure was developed, and undifferentiated human cord blood mesenchymal stem cells, in vitro hepato-genically differentiated mesenchymal stem cells, or freshly isolated rat hepatocytes were transplanted into a CCL4 cirrhotic experimental model. Animals were killed 3 months after transplant, and liver functions and histopathology were assessed. Compared with the cirrhotic control group, the 3 cell-treated groups showed improved alanine aminotransferase, aspartate aminotransferase, albumin, and bilirubin levels, with best results shown in the hepatocyte-treated group. Histopathologic examination of the treated groups showed improved fibrosis, with best results obtained in the undifferentiated mesenchymal stem cell-treated group. Both adult hepatocytes and cord blood mesenchymal stem cells proved to be promising candidates for cell-based therapy in liver regeneration on an experimental level. Improved liver function was evident in the hepatocyte-treated group, and fibrosis control was more evident in the undifferentiated mesenchymal stem cell-treated group.

The human Vδ2+ T-cell compartment comprises distinct innate-like Vγ9+ and adaptive Vγ9- subsets.

PubMed

Davey, Martin S; Willcox, Carrie R; Hunter, Stuart; Kasatskaya, Sofya A; Remmerswaal, Ester B M; Salim, Mahboob; Mohammed, Fiyaz; Bemelman, Frederike J; Chudakov, Dmitriy M; Oo, Ye H; Willcox, Benjamin E

2018-05-02

Vδ2 + T cells form the predominant human γδ T-cell population in peripheral blood and mediate T-cell receptor (TCR)-dependent anti-microbial and anti-tumour immunity. Here we show that the Vδ2 + compartment comprises both innate-like and adaptive subsets. Vγ9 + Vδ2 + T cells display semi-invariant TCR repertoires, featuring public Vγ9 TCR sequences equivalent in cord and adult blood. By contrast, we also identify a separate, Vγ9 - Vδ2 + T-cell subset that typically has a CD27 hi CCR7 + CD28 + IL-7Rα + naive-like phenotype and a diverse TCR repertoire, however in response to viral infection, undergoes clonal expansion and differentiation to a CD27 lo CD45RA + CX 3 CR1 + granzymeA/B + effector phenotype. Consistent with a function in solid tissue immunosurveillance, we detect human intrahepatic Vγ9 - Vδ2 + T cells featuring dominant clonal expansions and an effector phenotype. These findings redefine human γδ T-cell subsets by delineating the Vδ2 + T-cell compartment into innate-like (Vγ9 + ) and adaptive (Vγ9 - ) subsets, which have distinct functions in microbial immunosurveillance.

Short-term effects of stored homologous red blood cell transfusion on cardiorespiratory function and inflammation: an experimental study in a hypovolemia model

PubMed Central

Biagini, S.; Dale, C.S.; Real, J.M.; Moreira, E.S.; Carvalho, C.R.R.; Schettino, G.P.P.; Wendel, S.; Azevedo, L.C.P.

2017-01-01

The pathophysiological mechanisms associated with the effects of red blood cell (RBC) transfusion on cardiopulmonary function and inflammation are unclear. We developed an experimental model of homologous 14-days stored RBC transfusion in hypovolemic swine to evaluate the short-term effects of transfusion on cardiopulmonary system and inflammation. Sixteen healthy male anesthetized swine (68±3.3 kg) were submitted to controlled hemorrhage (25% of blood volume). Two units of non-filtered RBC from each animal were stored under blood bank conditions for 14 days. After 30 min of hypovolemia, the control group (n=8) received an infusion of lactated Ringer's solution (three times the removed volume). The transfusion group (n=8) received two units of homologous 14-days stored RBC and lactated Ringer's solution in a volume that was three times the difference between blood removed and blood transfusion infused. Both groups were followed up for 6 h after resuscitation with collection of hemodynamic and respiratory data. Cytokines and RNA expression were measured in plasma and lung tissue. Stored RBC transfusion significantly increased mixed oxygen venous saturation and arterial oxygen content. Transfusion was not associated with alterations on pulmonary function. Pulmonary concentrations of cytokines were not different between groups. Gene expression for lung cytokines demonstrated a 2-fold increase in mRNA level for inducible nitric oxide synthase and a 0.5-fold decrease in mRNA content for IL-21 in the transfused group. Thus, stored homologous RBC transfusion in a hypovolemia model improved cardiovascular parameters but did not induce significant effects on microcirculation, pulmonary inflammation and respiratory function up to 6 h after transfusion. PMID:29185590

Divergent response profile in activated cord blood T cells from first-born child implies birth-order-associated in utero immune programming.

PubMed

Kragh, M; Larsen, J M; Thysen, A H; Rasmussen, M A; Wolsk, H M; Bisgaard, H; Brix, S

2016-03-01

First-born children are at higher risk of developing a range of immune-mediated diseases. The underlying mechanism of 'birth-order effects' on disease risk is largely unknown, but in utero programming of the child's immune system may play a role. We studied the association between birth order and the functional response of stimulated cord blood T cells. Purified cord blood T cells were polyclonally activated with anti-CD3-/anti-CD28-coated beads in a subgroup of 28 children enrolled in the COPSAC2010 birth cohort. Expression levels of seven activation markers on helper and cytotoxic T cells as well as the percentage of CD4(+) CD25(+) T cells were assessed by flow cytometry. Production of IFN-γ, TNF-α, IL-17, IL-4, IL-5, IL-13, and IL-10 was measured in the supernatants. IL-10 secretion (P = 0.007) and CD25 expression on CD4(+) helper T cells (P = 0.0003) in the activated cord blood T cells were selectively reduced in first-born children, while the percentage of circulating CD4(+) CD25(+) cord blood T cells was independent of birth order. First-born infants display a reduced anti-inflammatory profile in T cells at birth. This possible in utero 'birth-order' T-cell programming may contribute to later development of immune-mediated diseases by increasing overall immune reactivity in first-born children as compared to younger siblings. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Mobilization of hematopoietic stem cells with highest self-renewal by G-CSF precedes clonogenic cell mobilization peak.

PubMed

Winkler, Ingrid G; Wiercinska, Eliza; Barbier, Valerie; Nowlan, Bianca; Bonig, Halvard; Levesque, Jean-Pierre

2016-04-01

Harvest of granulocyte colony-stimulating factor (G-CSF)-mobilized hematopoietic stem cells (HSCs) begins at day 5 of G-CSF administration, when most donors have achieved maximal mobilization. This is based on surrogate markers for HSC mobilization, such as CD34(+) cells and colony-forming activity in blood. However, CD34(+) cells or colony-forming units in culture (CFU-C) are heterogeneous cell populations with hugely divergent long-term repopulation potential on transplantation. HSC behavior is influenced by the vascular bed in the vicinity of which they reside. We hypothesized that G-CSF may mobilize sequentially cells proximal and more distal to bone marrow venous sinuses where HSCs enter the blood. We addressed this question with functional serial transplantation assays using blood and bone marrow after specific time points of G-CSF treatment in mice. We found that in mice, blood collected after only 48 hours of G-CSF administration was as enriched in serially reconstituting HSCs as blood collected at 5 days of G-CSF treatment. Similarly, mobilized Lin(-)CD34(+) cells were relatively enriched in more primitive Lin(-)CD34(+)CD38(-) cells at day 2 of G-CSF treatment compared with later points in half of human donors tested (n = 6). This suggests that in both humans and mice, hematopoietic progenitor and stem cells do not mobilize uniformly according to their maturation stage, with most potent HSCs mobilizing as early as day 2 of G-CSF. Copyright © 2016 ISEH - International Society for Experimental Hematology. Published by Elsevier Inc. All rights reserved.

Glycoprotein Ibα receptor instability is associated with loss of quality in platelets produced in culture.

PubMed

Robert, Amélie; Boyer, Lucie; Pineault, Nicolas

2011-03-01

The development of culture processes for hematopoietic progenitors could lead to the development of a complementary source of platelets for therapeutic purposes. However, functional characterization of culture-derived platelets remains limited, which raises some uncertainties about the quality of platelets produced in vitro. The aim of this study was to define the proportion of functional platelets produced in cord blood CD34+ cell cultures. Toward this, the morphological and functional properties of culture-derived platelet-like particles (PLPs) were critically compared to that of blood platelets. Flow cytometry combined with transmission electron microscopy analyses revealed that PLPs formed a more heterogeneous population of platelets at a different stage of maturation than blood platelets. The majority of PLPs harbored the fibrinogen receptor αIIbβ3, but a significant proportion failed to maintain glycoprotein (GP)Ibα surface expression, a component of the vWF receptor essential for platelet functions. Importantly, GPIbα extracellular expression correlated closely with platelet function, as the GPIIb+ GPIbα+ PLP subfraction responded normally to agonist stimulation as evidenced by α-granule release, adhesion, spreading, and aggregation. In contrast, the GPIIb+ GPIbα⁻ subfraction was unresponsive in most functional assays and appeared to be metabolically inactive. The present study confirms that functional platelets can be generated in cord blood CD34+ cell cultures, though these are highly susceptible to ectodomain shedding of receptors associated with loss of function. Optimization of culture conditions to prevent these deleterious effects and to homogenize PLPs is necessary to improve the quality and yields of culture-derived platelets before they can be recognized as a suitable complementary source for therapeutic purposes.

Development of a photon-cell interactive monte carlo simulation for non-invasive measurement of blood glucose level by Raman spectroscopy.

PubMed

Sakota, Daisuke; Kosaka, Ryo; Nishida, Masahiro; Maruyama, Osamu

2015-01-01

Turbidity variation is one of the major limitations in Raman spectroscopy for quantifying blood components, such as glucose, non-invasively. To overcome this limitation, we have developed a Raman scattering simulation using a photon-cell interactive Monte Carlo (pciMC) model that tracks photon migration in both the extra- and intracellular spaces without relying on the macroscopic scattering phase function and anisotropy factor. The interaction of photons at the plasma-cell boundary of randomly oriented three-dimensionally biconcave red blood cells (RBCs) is modeled using geometric optics. The validity of the developed pciMCRaman was investigated by comparing simulation and experimental results of Raman spectroscopy of glucose level in a bovine blood sample. The scattering of the excitation laser at a wavelength of 785 nm was simulated considering the changes in the refractive index of the extracellular solution. Based on the excitation laser photon distribution within the blood, the Raman photon derived from the hemoglobin and glucose molecule at the Raman shift of 1140 cm(-1) = 862 nm was generated, and the photons reaching the detection area were counted. The simulation and experimental results showed good correlation. It is speculated that pciMCRaman can provide information about the ability and limitations of the measurement of blood glucose level.

Extraction of superoxide dismutase, catalase, and carbonic anhydrase from stroma-free red blood cell hemolysate for the preparation of the nanobiotechnological complex of polyhemoglobin-superoxide dismutase-catalase-carbonic anhydrase.

PubMed

Guo, C; Gynn, M; Chang, T M S

2015-06-01

We report a novel method to simultaneously extract superoxide dismutase (SOD), catalase (CAT), and carbonic anhydrase (CA) from the same sample of red blood cells (RBCs). This avoids the need to use expensive commercial enzymes, thus enabling a cost-effective process for large-scale production of a nanobiotechnological polyHb-SOD-CAT-CA complex, with enhancement of all three red blood cell functions. An optimal concentration of phosphate buffer for ethanol-chloroform treatment results in good recovery of CAT, SOD, and CA after extraction. Different concentrations of the enzymes can be used to enhance the activity of polyHb-SOD-CAT-CA to 2, 4, or 6 times that of RBC.

Dysregulation of chemokine receptor expression and function in leukocytes from ALS patients.

PubMed

Perner, Caroline; Perner, Florian; Stubendorff, Beatrice; Förster, Martin; Witte, Otto W; Heidel, Florian H; Prell, Tino; Grosskreutz, Julian

2018-03-28

Amyotrophic lateral sclerosis (ALS) is rapidly progressive adult-onset motor neuron disease characterized by the neurodegeneration of both upper and lower motor neurons in the cortex and the spinal cord; the majority of patients succumb to respiratory failure. Although the etiology is not yet fully understood, there is compelling evidence that ALS is a multi-systemic disorder, with peripheral inflammation critically contributing to the disease process. However, the full extent and nature of this immunological dysregulation remains to be established, particularly within circulating blood cells. Therefore, the aim of the present study was to identify dysregulated inflammatory molecules in peripheral blood cells of ALS patients and analyze for functional consequences of the observed findings. To this end, we employed flow cytometry-based screening to quantify the surface expression of major chemokine receptors and integrins. A significantly increased expression of CXCR3, CXCR4, CCL2, and CCL5 was observed on T cells in ALS patients compared to healthy controls. Intriguingly, the expression was even more pronounced in patients with a slow progressive phenotype. To further investigate the functional consequences of this altered surface expression, we used a modified Boyden chamber assay to measure chemotaxis in ALS patient-derived lymphocytes. Interestingly, chemoattraction with the CXCR3-Ligand IP10 led to upregulated migratory behavior of ALS lymphocytes compared to healthy controls. Taken together, our data provides evidence for a functional dysregulation of IP10-directed chemotaxis in peripheral blood cells in ALS patients. However, whether the chemokine itself or its receptor CXCR3, or both, could serve as potential therapeutic targets in ALS requires further investigations.

Therapeutics incorporating blood constituents.

PubMed

Charoenphol, Phapanin; Oswalt, Katie; Bishop, Corey J

2018-04-05

Blood deficiency and dysfunctionality can result in adverse events, which can primarily be treated by transfusion of blood or the re-introduction of properly functioning sub-components. Blood constituents can be engineered on the sub-cellular (i.e., DNA recombinant technology) and cellular level (i.e., cellular hitchhiking for drug delivery) for supplementing and enhancing therapeutic efficacy, in addition to rectifying dysfunctioning mechanisms (i.e., clotting). Herein, we report the progress of blood-based therapeutics, with an emphasis on recent applications of blood transfusion, blood cell-based therapies and biomimetic carriers. Clinically translated technologies and commercial products of blood-based therapeutics are subsequently highlighted and perspectives on challenges and future prospects are discussed. Blood-based therapeutics is a burgeoning field and has advanced considerably in recent years. Blood and its constituents, with and without modification (i.e., combinatorial), have been utilized in a broad spectrum of pre-clinical and clinically-translated treatments. This review article summarizes the most up-to-date progress of blood-based therapeutics in the following contexts: synthetic blood substitutes, acellular/non-recombinant therapies, cell-based therapies, and therapeutic sub-components. The article subsequently discusses clinically-translated technologies and future prospects thereof. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Reprogramming mouse fibroblasts into engraftable myeloerythroid and lymphoid progenitors

PubMed Central

Cheng, Hui; Ang, Heather Yin-Kuan; A. EL Farran, Chadi; Li, Pin; Fang, Hai Tong; Liu, Tong Ming; Kong, Say Li; Chin, Michael Lingzi; Ling, Wei Yin; Lim, Edwin Kok Hao; Li, Hu; Huber, Tara; Loh, Kyle M.; Loh, Yuin-Han; Lim, Bing

2016-01-01

Recent efforts have attempted to convert non-blood cells into hematopoietic stem cells (HSCs) with the goal of generating blood lineages de novo. Here we show that hematopoietic transcription factors Scl, Lmo2, Runx1 and Bmi1 can convert a developmentally distant lineage (fibroblasts) into ‘induced hematopoietic progenitors' (iHPs). Functionally, iHPs generate acetylcholinesterase+ megakaryocytes and phagocytic myeloid cells in vitro and can also engraft immunodeficient mice, generating myeloerythoid and B-lymphoid cells for up to 4 months in vivo. Molecularly, iHPs transcriptionally resemble native Kit+ hematopoietic progenitors. Mechanistically, reprogramming factor Lmo2 implements a hematopoietic programme in fibroblasts by rapidly binding to and upregulating the Hhex and Gfi1 genes within days. Moreover the reprogramming transcription factors also require extracellular BMP and MEK signalling to cooperatively effectuate reprogramming. Thus, the transcription factors that orchestrate embryonic hematopoiesis can artificially reconstitute this programme in developmentally distant fibroblasts, converting them into engraftable blood progenitors. PMID:27869129

A mesoscopic simulation on distributions of red blood cells in a bifurcating channel

NASA Astrophysics Data System (ADS)

Inoue, Yasuhiro; Takagi, Shu; Matsumoto, Yoichiro

2004-11-01

Transports of red blood cells (RBCs) or particles in bifurcated channels have been attracting renewed interest since the advent of concepts of MEMS for sorting, analyzing, and removing cells or particles from sample medium. In this talk, we present a result on a transport of red blood cells (RBCs) in a bifurcating channel studied by using a mesoscale simulation technique of immiscible droplets, where RBCs have been modeled as immiscible droplets. The distribution of RBCs is represented by the fractional RBC flux into two daughters as a function of volumetric flow ratio between the daughters. The data obtained in our simulations are examined with a theoretical prediction, in which, we assume an exponential distribution for positions of RBCs in the mother channel. The theoretical predictions show a good agreement with simulation results. A non-uniform distribution of RBCs in the mother channel affects disproportional separation of RBC flux at a bifurcation.

Redox-dependent impairment of vascular function in sickle cell disease.

PubMed

Aslan, Mutay; Freeman, Bruce A

2007-12-01

The vascular pathophysiology of sickle cell disease (SCD) is influenced by many factors, including adhesiveness of red and white blood cells to endothelium, increased coagulation, and homeostatic perturbation. The vascular endothelium is central to disease pathogenesis because it displays adhesion molecules for blood cells, balances procoagulant and anticoagulant properties of the vessel wall, and regulates vascular homeostasis by synthesizing vasoconstricting and vasodilating substances. The occurrence of intermittent vascular occlusion in SCD leads to reperfusion injury associated with granulocyte accumulation and enhanced production of reactive oxygen species. The participation of nitric oxide (NO) in oxidative reactions causes a reduction in NO bioavailability and contributes to vascular dysfunction in SCD. Therapeutic strategies designed to counteract endothelial, inflammatory, and oxidative abnormalities may reduce the frequency of hospitalization and blood transfusion, the incidence of pain, and the occurrence of acute chest syndrome and pulmonary hypertension in patients with SCD.

Mapping the distribution of specific antibody interaction forces on individual red blood cells

NASA Astrophysics Data System (ADS)

Yeow, Natasha; Tabor, Rico F.; Garnier, Gil

2017-02-01

Current blood typing methods rely on the agglutination of red blood cells (RBCs) to macroscopically indicate a positive result. An indirect agglutination mechanism is required when blood typing with IgG forms of antibodies. To date, the interaction forces between anti-IgG and IgG antibodies have been poorly quantified, and blood group related antigens have never been quantified with the atomic force microscope (AFM). Instead, the total intensity resulting from fluorescent-tagged antibodies adsorbed on RBC has been measured to calculate an average antigen density on a series of RBCs. In this study we mapped specific antibody interaction forces on the RBC surface. AFM cantilever tips functionalized with anti-IgG were used to probe RBCs incubated with specific IgG antibodies. This work provides unique insight into antibody-antigen interactions in their native cell-bound location, and crucially, on a per-cell basis rather than an ensemble average set of properties. Force profiles obtained from the AFM directly provide not only the anti-IgG - IgG antibody interaction force, but also the spatial distribution and density of antigens over a single cell. This new understanding might be translated into the development of very selective and quantitative interactions that underpin the action of drugs in the treatment of frontier illnesses.

Effects of Electromagnetic Fields on Automated Blood Cell Measurements.

PubMed

Vagdatli, Eleni; Konstandinidou, Vasiliki; Adrianakis, Nikolaos; Tsikopoulos, Ioannis; Tsikopoulos, Alexios; Mitsopoulou, Kyriaki

2014-08-01

The aim of this study is to investigate whether the electromagnetic fields associated with mobile phones and/or laptops interfere with blood cell counts of hematology analyzers. Random blood samples were analyzed on an Aperture Impedance hematology analyzer. The analysis was performed in four ways: (A) without the presence of any mobile phone or portable computer in use, (B) with mobile phones in use (B1: one mobile, B4: four mobiles), (C) with portable computers (laptops) in use (C1: one laptop, C3: three laptops), and (D) with four mobile phones and three laptops in use simultaneously. The results obtained demonstrated a statistically significant decrease in neutrophil, erythrocyte, and platelet count and an increase in lymphocyte count, mean corpuscular volume, and red blood cell distribution width, notably in the B4 group. Despite this statistical significance, in clinical practice, only the red blood cell reduction could be taken into account, as the mean difference between the A and B4 group was 60,000 cells/µL. In group D, the analyzer gave odd results after 11 measurements and finally stopped working. The combined and multiple use of mobile phones and computers affects the function of hematology analyzers, leading to false results. Consequently, the use of such electronic devices must be avoided. © 2014 Society for Laboratory Automation and Screening.

Single-cell transcriptional analysis of normal, aberrant, and malignant hematopoiesis in zebrafish.

PubMed

Moore, Finola E; Garcia, Elaine G; Lobbardi, Riadh; Jain, Esha; Tang, Qin; Moore, John C; Cortes, Mauricio; Molodtsov, Aleksey; Kasheta, Melissa; Luo, Christina C; Garcia, Amaris J; Mylvaganam, Ravi; Yoder, Jeffrey A; Blackburn, Jessica S; Sadreyev, Ruslan I; Ceol, Craig J; North, Trista E; Langenau, David M

2016-05-30

Hematopoiesis culminates in the production of functionally heterogeneous blood cell types. In zebrafish, the lack of cell surface antibodies has compelled researchers to use fluorescent transgenic reporter lines to label specific blood cell fractions. However, these approaches are limited by the availability of transgenic lines and fluorescent protein combinations that can be distinguished. Here, we have transcriptionally profiled single hematopoietic cells from zebrafish to define erythroid, myeloid, B, and T cell lineages. We also used our approach to identify hematopoietic stem and progenitor cells and a novel NK-lysin 4(+) cell type, representing a putative cytotoxic T/NK cell. Our platform also quantified hematopoietic defects in rag2(E450fs) mutant fish and showed that these fish have reduced T cells with a subsequent expansion of NK-lysin 4(+) cells and myeloid cells. These data suggest compensatory regulation of the innate immune system in rag2(E450fs) mutant zebrafish. Finally, analysis of Myc-induced T cell acute lymphoblastic leukemia showed that cells are arrested at the CD4(+)/CD8(+) cortical thymocyte stage and that a subset of leukemia cells inappropriately reexpress stem cell genes, including bmi1 and cmyb In total, our experiments provide new tools and biological insights into single-cell heterogeneity found in zebrafish blood and leukemia. © 2016 Moore et al.

Single-cell transcriptional analysis of normal, aberrant, and malignant hematopoiesis in zebrafish

PubMed Central

Garcia, Elaine G.; Lobbardi, Riadh; Jain, Esha; Tang, Qin; Moore, John C.; Cortes, Mauricio; Molodtsov, Aleksey; Kasheta, Melissa; Luo, Christina C.; Garcia, Amaris J.; Mylvaganam, Ravi; Yoder, Jeffrey A.; Blackburn, Jessica S.; Sadreyev, Ruslan I.; Ceol, Craig J.; North, Trista E.

2016-01-01

Hematopoiesis culminates in the production of functionally heterogeneous blood cell types. In zebrafish, the lack of cell surface antibodies has compelled researchers to use fluorescent transgenic reporter lines to label specific blood cell fractions. However, these approaches are limited by the availability of transgenic lines and fluorescent protein combinations that can be distinguished. Here, we have transcriptionally profiled single hematopoietic cells from zebrafish to define erythroid, myeloid, B, and T cell lineages. We also used our approach to identify hematopoietic stem and progenitor cells and a novel NK-lysin 4+ cell type, representing a putative cytotoxic T/NK cell. Our platform also quantified hematopoietic defects in rag2E450fs mutant fish and showed that these fish have reduced T cells with a subsequent expansion of NK-lysin 4+ cells and myeloid cells. These data suggest compensatory regulation of the innate immune system in rag2E450fs mutant zebrafish. Finally, analysis of Myc-induced T cell acute lymphoblastic leukemia showed that cells are arrested at the CD4+/CD8+ cortical thymocyte stage and that a subset of leukemia cells inappropriately reexpress stem cell genes, including bmi1 and cmyb. In total, our experiments provide new tools and biological insights into single-cell heterogeneity found in zebrafish blood and leukemia. PMID:27139488

Hematopoietic Stem Cells in Neonates: Any Differences between Very Preterm and Term Neonates?

PubMed Central

Wisgrill, Lukas; Schüller, Simone; Bammer, Markus; Berger, Angelika; Pollak, Arnold; Radke, Teja Falk; Kögler, Gesine; Spittler, Andreas; Helmer, Hanns; Husslein, Peter; Gortner, Ludwig

2014-01-01

Background In the last decades, human full-term cord blood was extensively investigated as a potential source of hematopoietic stem and progenitor cells (HSPCs). Despite the growing interest of regenerative therapies in preterm neonates, only little is known about the biological function of HSPCs from early preterm neonates under different perinatal conditions. Therefore, we investigated the concentration, the clonogenic capacity and the influence of obstetric/perinatal complications and maternal history on HSPC subsets in preterm and term cord blood. Methods CD34+ HSPC subsets in UCB of 30 preterm and 30 term infants were evaluated by flow cytometry. Clonogenic assays suitable for detection of the proliferative potential of HSPCs were conducted. Furthermore, we analyzed the clonogenic potential of isolated HSPCs according to the stem cell marker CD133 and aldehyde dehydrogenase (ALDH) activity. Results Preterm cord blood contained a significantly higher concentration of circulating CD34+ HSPCs, especially primitive progenitors, than term cord blood. The clonogenic capacity of HSPCs was enhanced in preterm cord blood. Using univariate analysis, the number and clonogenic potential of circulating UCB HSPCs was influenced by gestational age, birth weight and maternal age. Multivariate analysis showed that main factors that significantly influenced the HSPC count were maternal age, gestational age and white blood cell count. Further, only gestational age significantly influenced the clonogenic potential of UCB HSPCs. Finally, isolated CD34+/CD133+, CD34+/CD133– and ALDHhigh HSPC obtained from preterm cord blood showed a significantly higher clonogenic potential compared to term cord blood. Conclusion We demonstrate that preterm cord blood exhibits a higher HSPC concentration and increased clonogenic capacity compared to term neonates. These data may imply an emerging use of HSPCs in autologous stem cell therapy in preterm neonates. PMID:25181353

[Immune dysfunction and cognitive deficit in stress and physiological aging. Part II: New approaches to cognitive disorder prevention and treatment ].

PubMed

Pukhal'skiĭ, A L; Shmarina, G V; Aleshkin, V A

2014-01-01

Long-term stress as well as physiological aging result in similar immunological and hormonal disturbances including hypothalamic-pituitary-adrenal) axis depletion, aberrant immune response (regulatory T-cells, Tregs, and T(h17)-lymphocyte accumulation) and decreased dehydroepian-drosterone synthesis both in the brain and in the adrenal glands. Since the main mechanisms of inflammation control, "prompt" (stress hormones) and "delayed" (Tregs), are broken, serum cytokine levels increase and become sufficient for blood-brain-barrier disruption. As a result peripheral cytokines penetrate into the brain where they begin to perform new functions. Structural and functional alterations of blood-brain-barrier as well as stress- (or age-) induced neuroinflammation promote influx of bone marrow derived dendritic cells and lymphocyte effectors into the brain parenchyma. Thereafter, mass intrusion ofpro-inflammatory mediators and immune cells having a lot of specific targets alters the brain work that we can observe both in humans and in animal experiments. The concept of stressful cognitive dysfunction, which is under consideration in this review, allows picking out several therapeutic targets: 1) reduction of excessive Treg accumulation; 2) supporting hypothalamic-pituitary-adrenal axis and inflammatory reaction attenuation; 3) recovery of dehydroepiandrosterone level; 4) improvement of blood-brain-barrier function.

Superiority of a functional leukocyte adhesiveness/aggregation test over the white blood cell count to discriminate between mild and significant inflammatory response in patients with acute bacterial infections.

PubMed

Rogowski, Ori; Rotstein, Rivka; Zeltzer, David; Misgav, Sarit; Justo, Daniel; Avitzour, Daniel; Mardi, Tamar; Serov, Jacob; Arber, Nadir; Berliner, Shlomo; Shapira, Itzhak

2002-01-01

Electronic cell counters may underestimate the white blood cell count (WBCC) in the presence of aggregated leukocytes. In the present study we focused on the possibility of using a functional, as opposed to an anatomic, count to circumvent this eventual underestimation. A model of bacterial infection was used because of the importance of leukocytosis in the physician's clinical decision-making process. There were 35 patients with low C-reactive protein (CRP) concentrations (0.5-4.9 mg/dL), 45 with intermediate (5-9.9 mg/dL), and 120 with relatively high (>10 mg/dL) CRP concentrations. A significant (P=0.008) difference was noted between the state of leukocyte adhesiveness/aggregation in the peripheral blood of individuals with low CRP concentrations (3.5%+/-4.3%) and those with high CRP concentrations (7.4%+/-8%), while there was no significant difference in the respective number of WBCs per cubic millimeter (cmm) (11,600 +/- 5,500 and 14,000 +/- 7,200, respectively). We raise the possibility that a functional test might be superior over an anatomic count in patients with acute bacterial infection and a significant acute phase response. Copyright 2002 Wiley-Liss, Inc.

Physiological responses and evaluation of effects of BMI, smoking and drinking in high altitude acclimatization: a cohort study in Chinese Han young males.

PubMed

Peng, Qian-Qian; Basang, Zhuoma; Cui, Chao-Ying; Li, Lei; Qian, Ji; Gesang, Quzhen; Yang, La; La, Zong; De, Yang; Dawa, Puchi; Qu, Ni; Suo, Qu; Dan, Zhen; Xiao, Duoji; Wang, Xiao-Feng; Jin, Li

2013-01-01

High altitude acclimatization is a series of physiological responses taking places when subjects go to altitude. Many factors could influence these processes, such as altitude, ascending speed and individual characteristics. In this study, based on a repeated measurement design of three sequential measurements at baseline, acute phase and chronic phase, we evaluated the effect of BMI, smoking and drinking on a number of physiological responses in high altitude acclimatization by using mixed model and partial least square path model on a sample of 755 Han Chinese young males. We found that subjects with higher BMI responses were reluctant to hypoxia. The effect of smoking was not significant at acute phase. But at chronic phase, red blood cell volume increased less while respiratory function increased more for smoking subjects compared with nonsmokers. For drinking subjects, red blood cell volume increased less than nondrinkers at both acute and chronic phases, while blood pressures increased more than nondrinkers at acute phase and respiratory function, red blood cell volume and oxygen saturation increased more than nondrinkers at chronic phase. The heavy and long-term effect of smoking, drinking and other factors in high altitude acclimatization needed to be further studied.

Physiological Responses and Evaluation of Effects of BMI, Smoking and Drinking in High Altitude Acclimatization: A Cohort Study in Chinese Han Young Males

PubMed Central

Cui, Chao-ying; Li, Lei; Qian, Ji; Gesang, Quzhen; Yang, La; La, Zong; De, Yang; Dawa, Puchi; Qu, Ni; Suo, Qu; Dan, Zhen; Xiao, Duoji; Wang, Xiao-feng; Jin, Li

2013-01-01

High altitude acclimatization is a series of physiological responses taking places when subjects go to altitude. Many factors could influence these processes, such as altitude, ascending speed and individual characteristics. In this study, based on a repeated measurement design of three sequential measurements at baseline, acute phase and chronic phase, we evaluated the effect of BMI, smoking and drinking on a number of physiological responses in high altitude acclimatization by using mixed model and partial least square path model on a sample of 755 Han Chinese young males. We found that subjects with higher BMI responses were reluctant to hypoxia. The effect of smoking was not significant at acute phase. But at chronic phase, red blood cell volume increased less while respiratory function increased more for smoking subjects compared with nonsmokers. For drinking subjects, red blood cell volume increased less than nondrinkers at both acute and chronic phases, while blood pressures increased more than nondrinkers at acute phase and respiratory function, red blood cell volume and oxygen saturation increased more than nondrinkers at chronic phase. The heavy and long-term effect of smoking, drinking and other factors in high altitude acclimatization needed to be further studied. PMID:24260204

Effects of PEG-PLA-nano Artificial Cells Containing Hemoglobin on Kidney Function and Renal Histology in Rats

PubMed Central

Liu, Zun Chang; Chang, Thomas M.S.

2012-01-01

This study is to investigate the long-term effects of PEG-PLA nano artificial cells containing hemoglobin (NanoRBC) on renal function and renal histology after 1/3 blood volume top loading in rats. The experimental rats received one of the following infusions: NanoRBC in Ringer lactate, Ringer lactate, stroma-free hemoglobin (SFHB), polyhemoglobin (PolyHb), autologous rat whole blood (rat RBC). Blood samples were taken before infusions and on days 1, 7 and 21 after infusions for biochemistry analysis. Rats were sacrificed on day 21 after infusions and kidneys were excised for histology examination. Infusion of SFHB induced significant decrease in renal function damage evidenced by elevated serum urea, creatinine and uric acid throughout the 21 days. Kidney histology in SFHb infusion group revealed focal tubular necrosis and intraluminal cellular debris in the proximal tubules, whereas the glomeruli were not observed damaged. In all the other groups, NanoRBC, PolyHb, Ringer lactate and rat RBC, there were no abnormalities in renal biochemistry or histology. In conclusion, injection of NanoRBC did not have adverse effects on renal function nor renal histology. PMID:18979292

Role of the frequency of blood CD4{sup +} CXCR5{sup +} CCR6{sup +} T cells in autoimmunity in patients with Sjoegren's syndrome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Xue-yi; Wu, Zhen-biao; Ding, Jin

2012-06-01

Highlights: Black-Right-Pointing-Pointer The frequency of CD4{sup +} CXCR5{sup +} CCR6{sup +} T cells increased in pSS patients and positively correlated with autoantibodies in the blood. Black-Right-Pointing-Pointer CD4{sup +} CXCR5{sup +} CCR6{sup +} T cells in blood invariably coexpressed PD-1, ICOS, CD40L, Bcl-6 and secreted IL-21 after stimulated by PHA. Black-Right-Pointing-Pointer CD4{sup +} CXCR5{sup +} CCR6{sup +} Tfh cells in blood may be suitable biomarkers for the evaluation of the active immune stage of pSS patients. -- Abstract: The blood CD4{sup +} CXCR5{sup +} T cells, known as 'circulating' Tfh, have been shown to efficiently induce naieve B cells to producemore » immunoglobulin. They play an important role in certain autoimmune diseases. In the present study, we show for the first time that the frequency of CD4{sup +} CXCR5{sup +} T cells is increased in pSS patients and positively correlated with autoantibodies in the blood. The concentration of Th17-like subsets (CD4{sup +} CXCR5{sup +} CCR6{sup +}) in pSS patients was found to be significantly higher than in healthy controls. Functional assays showed that activated Th17-like subtypes in the blood display the key features of Tfh cells, including invariably coexpressed PD-1, ICOS, CD40L and IL-21. Th17 subsets were found to highly express Bcl-6 protein and Th1 and Th2 were not. Bcl-6 is believed to be a master transforming factor for Tfh cell differentiation and facilitate B cell proliferation and somatic hypermutation within the germinal center. These data indicate that Th17 subsets of CD4{sup +} CXCR5{sup +} T cells in the blood may participate in the antibody-related immune responses and that high frequency of CD4{sup +} CXCR5{sup +} CCR6{sup +} Tfh cells in blood may be suitable biomarkers for the evaluation of the active immune stage of pSS patients. It might provide insights into the pathogenesis and perhaps help researchers identify novel therapeutic targets for pSS.« less

Phenotype and function of CD209+ bovine blood dendritic cells, monocyte-derived-dendritic cells and monocyte-derived macrophages

USDA-ARS?s Scientific Manuscript database

Phylogenic comparisons of the mononuclear phagocyte system (MPS) of humans and mice demonstrate phenotypic divergence of dendritic cell (DC) subsets that play similar roles in innate and adaptive immunity. Although differing in phenotype, DC can be classified into four groups according to ontogeny a...

Blood-Feeding Induces Reversible Functional Changes in Flight Muscle Mitochondria of Aedes aegypti Mosquito

PubMed Central

Gonçalves, Renata L. S.; Machado, Ana Carolina L.; Paiva-Silva, Gabriela O.; Sorgine, Marcos H. F.; Momoli, Marisa M.; Oliveira, Jose Henrique M.; Vannier-Santos, Marcos A.; Galina, Antonio; Oliveira, Pedro L.; Oliveira, Marcus F.

2009-01-01

Background Hematophagy poses a challenge to blood-feeding organisms since products of blood digestion can exert cellular deleterious effects. Mitochondria perform multiple roles in cell biology acting as the site of aerobic energy-transducing pathways, and also an important source of reactive oxygen species (ROS), modulating redox metabolism. Therefore, regulation of mitochondrial function should be relevant for hematophagous arthropods. Here, we investigated the effects of blood-feeding on flight muscle (FM) mitochondria from the mosquito Aedes aegypti, a vector of dengue and yellow fever. Methodology/Principal Findings Blood-feeding caused a reversible reduction in mitochondrial oxygen consumption, an event that was parallel to blood digestion. These changes were most intense at 24 h after blood meal (ABM), the peak of blood digestion, when oxygen consumption was inhibited by 68%. Cytochromes c and a+a 3 levels and cytochrome c oxidase activity of the electron transport chain were all reduced at 24 h ABM. Ultrastructural and molecular analyses of FM revealed that mitochondria fuse upon blood meal, a condition related to reduced ROS generation. Consistently, BF induced a reversible decrease in mitochondrial H2O2 formation during blood digestion, reaching their lowest values at 24 h ABM where a reduction of 51% was observed. Conclusion Blood-feeding triggers functional and structural changes in hematophagous insect mitochondria, which may represent an important adaptation to blood feeding. PMID:19924237

Notch3 is necessary for blood vessel integrity in the central nervous system.

PubMed

Henshall, Tanya L; Keller, Annika; He, Liqun; Johansson, Bengt R; Wallgard, Elisabet; Raschperger, Elisabeth; Mäe, Maarja Andaloussi; Jin, Shaobo; Betsholtz, Christer; Lendahl, Urban

2015-02-01

Vascular smooth muscle cells (VSMC) are important for contraction, blood flow distribution, and regulation of blood vessel diameter, but to what extent they contribute to the integrity of blood vessels and blood-brain barrier function is less well understood. In this report, we explored the impact of the loss of VSMC in the Notch3(-/-) mouse on blood vessel integrity in the central nervous system. Notch3(-/-) mice showed focal disruptions of the blood-brain barrier demonstrated by extravasation of tracers accompanied by fibrin deposition in the retinal vasculature. This blood-brain barrier leakage was accompanied by a regionalized and patchy loss of VSMC, with VSMC gaps predominantly in arterial resistance vessels of larger caliber. The loss of VSMC appeared to be caused by progressive degeneration of VSMC resulting in a gradual loss of VSMC marker expression and a progressive acquisition of an aberrant VSMC phenotype closer to the gaps, followed by enhanced apoptosis and cellular disintegration in the gaps. Arterial VSMC were the only mural cell type that was morphologically affected, despite Notch3 also being expressed in pericytes. Transcriptome analysis of isolated brain microvessels revealed gene expression changes in Notch3(-/-) mice consistent with loss of arterial VSMC and presumably secondary transcriptional changes were observed in endothelial genes, which may explain the compromised vascular integrity. We demonstrate that Notch3 is important for survival of VSMC, and reveal a critical role for Notch3 and VSMC in blood vessel integrity and blood-brain barrier function in the mammalian vasculature. © 2014 American Heart Association, Inc.

Haptoglobin Preserves Vascular Nitric Oxide Signaling during Hemolysis.

PubMed

Schaer, Christian A; Deuel, Jeremy W; Schildknecht, Daniela; Mahmoudi, Leila; Garcia-Rubio, Ines; Owczarek, Catherine; Schauer, Stefan; Kissner, Reinhard; Banerjee, Uddyalok; Palmer, Andre F; Spahn, Donat R; Irwin, David C; Vallelian, Florence; Buehler, Paul W; Schaer, Dominik J

2016-05-15

Hemolysis occurs not only in conditions such as sickle cell disease and malaria but also during transfusion of stored blood, extracorporeal circulation, and sepsis. Cell-free Hb depletes nitric oxide (NO) in the vasculature, causing vasoconstriction and eventually cardiovascular complications. We hypothesize that Hb-binding proteins may preserve vascular NO signaling during hemolysis. Characterization of an archetypical function by which Hb scavenger proteins could preserve NO signaling during hemolysis. We investigated NO reaction kinetics, effects on arterial NO signaling, and tissue distribution of cell-free Hb and its scavenger protein complexes. Extravascular translocation of cell-free Hb into interstitial spaces, including the vascular smooth muscle cell layer of rat and pig coronary arteries, promotes vascular NO resistance. This critical disease process is blocked by haptoglobin. Haptoglobin does not change NO dioxygenation rates of Hb; rather, the large size of the Hb:haptoglobin complex prevents Hb extravasation, which uncouples NO/Hb interaction and vasoconstriction. Size-selective compartmentalization of Hb functions as a substitute for red blood cells after hemolysis and preserves NO signaling in the vasculature. We found that evolutionarily and structurally unrelated Hb-binding proteins, such as PIT54 found in avian species, functionally converged with haptoglobin to protect NO signaling by sequestering cell-free Hb in large protein complexes. Sequential compartmentalization of Hb by erythrocytes and scavenger protein complexes is an archetypical mechanism, which may have supported coevolution of hemolysis and normal vascular function. Therapeutic supplementation of Hb scavengers may restore vascular NO signaling and attenuate disease complications in patients with hemolysis.

Cryopreservation of hematopoietic stem and progenitor cells amplified ex vivo from cord blood CD34+ cells.

PubMed

Duchez, Pascale; Chevaleyre, Jean; Brunet de la Grange, Philippe; Vlaski, Marija; Boiron, Jean-Michel; Wouters, Guy; Ivanovic, Zoran

2013-09-01

Our ex vivo expansion procedure starting from cord blood (CB) CD34+ cells enabled expansion of committed progenitors (CPs) without a negative impact on hematopoietic stem cells (HSCs) exhibiting both short- and long-term repopulating capacity. Upgraded to clinical scale (Macopharma HP01 in the presence of stem cell factor, FLT3-L [100 ng/mL each], granulocyte-colony-stimulating factor [10 ng/mL], and thrombopoietin [20 ng/mL]), it is being used for an ongoing clinical trial (adult allogeneic context) yielding promising preliminary results. Transplantation of ex vivo expanded CB cells is becoming a reality, while the issue of expanded cells' cryopreservation emerges as an option that allows the conservation of the product for transportation and future use. Here, we investigated whether it is possible to maintain the functional HSC and CP properties after freezing and thawing of expanded cells. We compared cryopreservation efficiency of the ex vivo expanded CB cells using the standard protocol (freezing solution human serum albumin (HSA)-dimethyl sulfoxide [DMSO]) with the newly designed protocol based on an enriched freezing solution (HP01-DMSO) with respect to the viability index, number of CD34+ and total cells, and recovery of CPs (colony-forming units) and HSCs (NOG/Scid/gamma-null mice engraftment). Cryopreservation and thawing of expanded CB cells using the "standard" procedure (HSA-DMSO) reduced recovery of the CPs (40%) and HSCs (drastically decreasing engraftment capacity). HP01-based protocol resulted in improvement of preservation of both CPs (>60%) and HSCs (nonaltered engraftment capacities). Functional maintenance of the expanded graft by cryopreservation is feasible in conditions compatible with human cell therapy requirements. © 2012 American Association of Blood Banks.

Effect of diastolic flow patterns on the function of the left ventricle

NASA Astrophysics Data System (ADS)

Seo, Jung Hee; Mittal, Rajat

2013-11-01

Direct numerical simulations are used to study the effect of intraventricular flow patterns on the pumping efficiency and the blood mixing and transport characteristics of the left ventricle. The simulations employ a geometric model of the left ventricle which is derived from contrast computed tomography. A variety of diastolic flow conditions are generated for a fixed ejection fraction in order to delineate the effect of flow patterns on ventricular performance. The simulations indicate that the effect of intraventricular blood flow pattern on the pumping power is physiologically insignificant. However, diastolic flow patterns have a noticeable effect on the blood mixing as well as the residence time of blood cells in the ventricle. The implications of these findings on ventricular function are discussed.

Quantitative blood group typing using surface plasmon resonance.

PubMed

Then, Whui Lyn; Aguilar, Marie-Isabel; Garnier, Gil

2015-11-15

The accurate and reliable typing of blood groups is essential prior to blood transfusion. While current blood typing methods are well established, results are subjective and heavily reliant on analysis by trained personnel. Techniques for quantifying blood group antibody-antigen interactions are also very limited. Many biosensing systems rely on surface plasmon resonance (SPR) detection to quantify biomolecular interactions. While SPR has been widely used for characterizing antibody-antigen interactions, measuring antibody interactions with whole cells is significantly less common. Previous studies utilized SPR for blood group antigen detection, however, showed poor regeneration causing loss of functionality after a single use. In this study, a fully regenerable, multi-functional platform for quantitative blood group typing via SPR detection is achieved by immobilizing anti-human IgG antibody to the sensor surface, which binds to the Fc region of human IgG antibodies. The surface becomes an interchangeable platform capable of quantifying the blood group interactions between red blood cells (RBCs) and IgG antibodies. As with indirect antiglobulin tests (IAT), which use IgG antibodies for detection, IgG antibodies are initially incubated with RBCs. This facilitates binding to the immobilized monolayer and allows for quantitative blood group detection. Using the D-antigen as an example, a clear distinction between positive (>500 RU) and negative (<100 RU) RBCs is achieved using anti-D IgG. Complete regeneration of the anti-human IgG surface is also successful, showing negligible degradation of the surface after more than 100 regenerations. This novel approach is validated with human-sourced whole blood samples to demonstrate an interesting alternative for quantitative blood grouping using SPR analysis. Crown Copyright © 2015. Published by Elsevier B.V. All rights reserved.

The effects of ILLLI on peripheral blood T lymphocytes subpopulation & NK cells in psoriasis treatment

NASA Astrophysics Data System (ADS)

Zhu, Jing; Nie, Fan

2005-07-01

Objective: To research the effects of Intravascular low level laser irradiation (ILLLI) on the immulogic function of cells in treatment of psoriasis. Method: 49 patients suffered from psoriasis were treated by Intravascular low level laser irradiation (laser output power: 4-5mw, 1 hour per day, a course of treatment is 10 days). We checked the function of T lymphocyte subgroup and NK cell in peripheral blood between pre and post treatment. Results: 1.The mean value of CD3+ in post treatment is higher. P<0.05. Significant difference is showed between pre and post treatment 2. The mean value of CD4+ in post treatment dropped slightly while the mean value of CD4/CD8, NK cell in post treatment increased little, nearly approach the mean value of natural person. 3.The mean value of CD4+,CD8+,NK cell which is under 30% increased the percent obviously after the treatment; The mean value of CD4+,CD8+ u higher than 30% obviously drop the percent, P#0.05 and <0.01. Related statistical analysis showed significant and much significant difference between pre and post treatment. Conclusions: The low level laser irradiation (ILLLI) in treatment of psoriasis has bidirectional ajustive effect which can balance the immulogic function of cell.

Postinfarction Functional Recovery Driven by a Three-Dimensional Engineered Fibrin Patch Composed of Human Umbilical Cord Blood-Derived Mesenchymal Stem Cells.

PubMed

Roura, Santiago; Soler-Botija, Carolina; Bagó, Juli R; Llucià-Valldeperas, Aida; Férnandez, Marco A; Gálvez-Montón, Carolina; Prat-Vidal, Cristina; Perea-Gil, Isaac; Blanco, Jerónimo; Bayes-Genis, Antoni

2015-08-01

Considerable research has been dedicated to restoring myocardial cell slippage and limiting ventricular remodeling after myocardial infarction (MI). We examined the ability of a three-dimensional (3D) engineered fibrin patch filled with human umbilical cord blood-derived mesenchymal stem cells (UCBMSCs) to induce recovery of cardiac function after MI. The UCBMSCs were modified to coexpress luciferase and fluorescent protein reporters, mixed with fibrin, and applied as an adhesive, viable construct (fibrin-cell patch) over the infarcted myocardium in mice (MI-UCBMSC group). The patch adhered well to the heart. Noninvasive bioluminescence imaging demonstrated early proliferation and differentiation of UCBMSCs within the construct in the postinfarct mice in the MI-UCBMSC group. The implanted cells also participated in the formation of new, functional microvasculature that connected the fibrin-cell patch to both the subjacent myocardial tissue and the host circulatory system. As revealed by echocardiography, the left ventricular ejection fraction and fractional shortening at sacrifice were improved in MI-UCBMSC mice and were markedly reduced in mice treated with fibrin alone and untreated postinfarction controls. In conclusion, a 3D engineered fibrin patch composed of UCBMSCs attenuated infarct-derived cardiac dysfunction when transplanted locally over a myocardial wound. ©AlphaMed Press.

Lipocalin 2 functions as a negative regulator of red blood cell production in an autocrine fashion.

PubMed

Miharada, Ken-ichi; Hiroyama, Takashi; Sudo, Kazuhiro; Nagasawa, Toshiro; Nakamura, Yukio

2005-11-01

Members of the lipocalin protein family are typically small, secreted proteins that possess a variety of functions. Although the physiological role of lipocalin 2 remains to be fully elucidated, a few pivotal functions have recently been reported, e.g., regulation of the apoptosis of leukocytes. Unexpectedly, lipocalin 2 is abundantly expressed in erythroid progenitor cells. An in vitro culture experiment demonstrated that lipocalin 2 induces apoptosis and inhibits differentiation of erythroid progenitor cells. During acute anemia the expression of lipocalin 2 was reduced in erythroid cells by a feedback system. Furthermore, injection of recombinant lipocalin 2 into mice suffering from acute anemia retarded the recovery of red blood cell (RBC) numbers, suggesting the importance of reduced expression of lipocalin 2 for the efficient recovery of RBC numbers. These results indicate that lipocalin 2 suppresses RBC production in an autocrine fashion. Hence, anemia arising from pathological conditions, such as chronic inflammation, might be partly due to increased levels of lipocalin 2 secreted from expanded leukocytes and/or macrophages. Also, anemia arising from malignancies might be partly due to the abundant secretion of lipocalin 2 from tumor cells. Thus, lipocalin 2 may represent an attractive therapeutic target for anemia under certain pathological conditions.

Mesenchymal Stem Cells Contribute to Improvement of Renal Function in a Canine Kidney Injury Model.

PubMed

Lee, Seung-Jun; Ryu, Min-Ok; Seo, Min-Soo; Park, Sang-Bum; Ahn, Jin-Ok; Han, Sei-Myoung; Kang, Kyung-Sun; Bhang, Dong-Ha; Youn, Hwa-Young

2017-01-01

The kidney excretes waste materials and regulates important metabolic functions, and renal disorders constitute a significant medical problem and can result in fatalities. In the present study, mesenchymal stem cells derived from canine umbilical cord blood (cUCB-MSCs) were isolated and evaluated for their ability to improve renal function in a canine model of acute kidney injury (AKI). The canine AKI model was developed by i.v. injection of cisplatin and gentamycin into 14 male beagle dogs. cUCB-MSCs were administered into the renal corticomedullary junction following AKI induction. Survival time, clinical signs, blood analysis and histological parameters were analyzed. The group treated with AKI plus cUCB-MSCs had decreased blood urea nitrogen and creatinine levels, and showed an extended life-span and improved histological manifestations. MSCs were detected around the tubules of these kidneys at the histological level. Taken together, our findings suggest that cUCB-MSCs could be an alternative therapeutic agent for canine AKI. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Cell Therapy From Bench to Bedside Translation in CNS Neurorestoratology Era

PubMed Central

Huang, Hongyun; Chen, Lin; Sanberg, Paul

2010-01-01

Recent advances in cell biology, neural injury and repair, and the progress towards development of neurorestorative interventions are the basis for increased optimism. Based on the complexity of the processes of demyelination and remyelination, degeneration and regeneration, damage and repair, functional loss and recovery, it would be expected that effective therapeutic approaches will require a combination of strategies encompassing neuroplasticity, immunomodulation, neuroprotection, neurorepair, neuroreplacement, and neuromodulation. Cell-based restorative treatment has become a new trend, and increasing data worldwide have strongly proven that it has a pivotal therapeutic value in CNS disease. Moreover, functional neurorestoration has been achieved to a certain extent in the CNS clinically. Up to now, the cells successfully used in preclinical experiments and/or clinical trial/treatment include fetal/embryonic brain and spinal cord tissue, stem cells (embryonic stem cells, neural stem/progenitor cells, hematopoietic stem cells, adipose-derived adult stem/precursor cells, skin-derived precursor, induced pluripotent stem cells), glial cells (Schwann cells, oligodendrocyte, olfactory ensheathing cells, astrocytes, microglia, tanycytes), neuronal cells (various phenotypic neurons and Purkinje cells), mesenchymal stromal cells originating from bone marrow, umbilical cord, and umbilical cord blood, epithelial cells derived from the layer of retina and amnion, menstrual blood-derived stem cells, Sertoli cells, and active macrophages, etc. Proof-of-concept indicates that we have now entered a new era in neurorestoratology. PMID:21359168

[Peripheral neuropathy and blood-nerve barrier].

PubMed

Kanda, Takashi

2009-11-01

It is important to know the cellular properties of endoneurial microvascular endothelial cells (PnMECs) and microvascular pericytes which constitute blood-nerve barrier (BNB), since this barrier structure in the peripheral nervous system (PNS) may play pivotal pathophysiological roles in various disorders of the PNS including inflammatory neuropathies (i.e. Guillain-Barré syndrome), vasculitic neuropathies, hereditary neuropathies and diabetic neuropathy. However, in contrast to blood-brain barrier (BBB), very few studies have been directed to BNB and no adequate cell lines originating from BNB had been launched. In our laboratory, we successfully established human immortalized cell lines originating from BNB using temperature-sensitive SV40 large T antigen and the cellular properties of human cell lines are presented in this paper. Human PnMEC cell line showed high transendothelial electrical resistance and expressed tight junction components and various types of influx as well as efflux transporters that have been reported to function at BBB. Human pericyte cell line also possessed tight junction proteins except claudin-5 and secrete various cytokines and growth factors including bFGF, VEGF, GDNF, NGF, BDNF and angiopoietin-1. Co-culture with pericytes or pericyte-conditioned media strengthend barrier properties of PnMEC, suggesting that in the PNS, peripheral nerve pericytes support the BNB function and play the same role of astrocytes in the BBB. Future accumulation of the knowledge concerning the cellular properties of BNB-forming cells will open the door to novel therapeutic strategies for intractable peripheral neuropathies.

Peripheral blood natural killer cells and mild thyroid abnormalities in women with reproductive failure

PubMed Central

Triggianese, P; Perricone, C; Conigliaro, P; Chimenti, MS; Perricone, R; De Carolis, C

2015-01-01

Abnormalities in peripheral blood natural killer (NK) cells have been reported in women with primary infertility and recurrent spontaneous abortion (RSA) and several studies have been presented to define cutoff values for abnormal peripheral blood NK cell levels in this context. Elevated levels of NK cells were observed in infertile/RSA women in the presence of thyroid autoimmunity (TAI), while no studies have been carried out, to date, on NK cells in infertile/RSA women with non-autoimmune thyroid diseases. The contribution of this study is two-fold: (1) the evaluation of peripheral blood NK cell levels in a cohort of infertile/RSA women, in order to confirm related data from the literature; and (2) the assessment of NK cell levels in the presence of both TAI and subclinical hypothyroidism (SCH) in order to explore the possibility that the association between NK cells and thyroid function is not only restricted to TAI but also to SCH. In a retrospective study, 259 age-matched women (primary infertility [n = 49], primary RSA [n = 145], and secondary RSA [n = 65]) were evaluated for CD56+CD16+NK cells by flow cytometry. Women were stratified according to thyroid status: TAI, SCH, and without thyroid diseases (ET). Fertile women (n = 45) were used as controls. Infertile/RSA women showed higher mean NK cell levels than controls. The cutoff value determining the abnormal NK cell levels resulted ⩾15% in all the groups of women. Among the infertile/RSA women, SCH resulted the most frequently associated thyroid disorder while no difference resulted in the prevalence of TAI and ET women between patients and controls. A higher prevalence of women with NK cell levels ⩾15% was observed in infertile/RSA women with SCH when compared to TAI/ET women. According to our data, NK cell assessment could be used as a diagnostic tool in women with reproductive failure and we suggest that the possible association between NK cell levels and thyroid function can be described not only in the presence of TAI but also in the presence of non-autoimmune thyroid disorders. PMID:26657164

Intra-renal delivery of mesenchymal stem cells attenuates myocardial injury after reversal of hypertension in porcine renovascular disease.

PubMed

Eirin, Alfonso; Zhu, Xiang-Yang; Ferguson, Christopher M; Riester, Scott M; van Wijnen, Andre J; Lerman, Amir; Lerman, Lilach O

2015-01-19

Percutaneous transluminal renal angioplasty (PTRA) fails to fully improve cardiac injury and dysfunction in patients with renovascular hypertension (RVH). Mesenchymal stem cells (MSCs) restore renal function, but their potential for attenuating cardiac injury after reversal of RVH has not been explored. We hypothesized that replenishment of MSCs during PTRA would improve cardiac function and oxygenation, and decrease myocardial injury in porcine RVH. Pigs were studied after 16 weeks of RVH, RVH treated 4 weeks earlier with PTRA with or without adjunct intra-renal delivery of MSC (10^6 cells), and controls. Cardiac structure, function (fast-computed tomography (CT)), and myocardial oxygenation (Blood-Oxygen-Level-Dependent- magnetic resonance imaging) were assessed in-vivo. Myocardial microvascular density (micro-CT) and myocardial injury were evaluated ex-vivo. Kidney venous and systemic blood levels of inflammatory markers were measured and their renal release calculated. PTRA normalized blood pressure, yet stenotic-kidney glomerular filtration rate, similarly blunted in RVH and RVH + PTRA, normalized only in PTRA + MSC-treated pigs. PTRA attenuated left ventricular remodeling, whereas myocardial oxygenation, subendocardial microvascular density, and diastolic function remained decreased in RVH + PTRA, but normalized in RVH + PTRA-MSC. Circulating isoprostane levels and renal release of inflammatory cytokines increased in RVH and RVH + PTRA, but normalized in RVH + PTRA-MSC, as did myocardial oxidative stress, inflammation, collagen deposition, and fibrosis. Intra-renal MSC delivery during PTRA preserved stenotic-kidney function, reduced systemic oxidative stress and inflammation, and thereby improved cardiac function, oxygenation, and myocardial injury four weeks after revascularization, suggesting a therapeutic potential for adjunctive MSC delivery to preserve cardiac function and structure after reversal of experimental RVH.

Innate immune responses in central nervous system inflammation.

PubMed

Finsen, Bente; Owens, Trevor

2011-12-01

In autoimmune diseases of the central nervous system (CNS), innate glial cell responses play a key role in determining the outcome of leukocyte infiltration. Access of leukocytes is controlled via complex interactions with glial components of the blood-brain barrier that include angiotensin II receptors on astrocytes and immunoregulatory mediators such as Type I interferons which regulate cellular traffic. Myeloid cells at the blood-brain barrier present antigen to T cells and influence cytokine effector function. Myelin-specific T cells interact with microglia and promote differentiation of oligodendrocyte precursor cells in response to axonal injury. These innate responses offer potential targets for immunomodulatory therapy. Copyright © 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

[Morphofunctional properties of the peripheral blood and bone marrow cells of rats following a flight on board the Kosmos-936 biosatellite].

PubMed

Kozinets, G I; Korol'kov, V I; Britvan, I I; Bykova, I A; Spitsyna, N E

1983-01-01

Morphofunctional properties of peripheral blood cells of Cosmos-936 rats were examined, using morphological, interferometric and electron microscopic techniques. As follows from the morphological data, immediately after recovery the weightless rats showed symptoms of a stress reaction which disappeared by R+3. The centrifuged rats exhibited less expressed symptoms of this sort. The percentage of bone marrow cell distribution was shifted towards enhanced myelopoiesis and diminished erythropoiesis. By the end of the readaptation period the ratio of bone marrow cell composition returned to normal. Interferometric and electron microscopic examinations did not reveal any irreversible changes in the structure and function of cells that may be caused by zero-g.

Beyond CD34+ cell dose: impact of method of peripheral blood hematopoietic stem cell mobilization (granulocyte-colony-stimulating factor [G-CSF], G-CSF plus plerixafor, or cyclophosphamide G-CSF/granulocyte-macrophage [GM]-CSF) on number of colony-forming unit-GM, engraftment, and Day +100 hematopoietic graft function.

PubMed

Alexander, Erin T; Towery, Jeanne A; Miller, Ashley N; Kramer, Cindy; Hogan, Kathy R; Squires, Jerry E; Stuart, Robert K; Costa, Luciano J

2011-09-01

The dose of CD34+ cells/kg in the mobilized peripheral blood product is the main determinant of neutrophil and platelet (PLT) engraftment after autologous hematopoietic stem cell transplantation (AHSCT). Whether the method of mobilization, namely, granulocyte-colony-stimulating factor (G-CSF) alone (G), G-CSF plus plerixafor (G+P), or cyclophosphamide + G/granulocyte-macrophage (GM)-CSF (Cy+G/GM), independently affects number of colony-forming unit (CFU)-GM, engraftment, and hematopoietic graft function is unknown. We used a database of AHSCT patients with multiple myeloma or lymphoma to identify three groups with different mobilization strategies receiving transplantation with similar CD34+ cell doses. Groups were compared in terms of CFU-GM, ratio of CFU-GM/CD34+, engraftment of neutrophils and PLTs, and hematopoietic graft function on Day +100. Ninety-six patients were included in the analysis, 26 G, 32 G+P, and 38 Cy+G/GM, with median cell doses of 4.21 × 10(6) , 4.11 × 10(6) , and 4.67 × 10(6) CD34+/kg, respectively (p = 0.433). There was no significant difference in number of CFU-GM between the three groups; however, the ratio of CFU-GM/CD34+ was significantly lower for G+P (p = 0.008). Median time for neutrophil engraftment was 13 days in G+P and 12 days in G and Cy+G/GM (p = 0.028), while PLT engraftment happened at a median of 14.5 days in G+P versus 12 days in G and 11 days in Cy+G/GM (p = 0.012). There was no difference in hematopoietic graft function at Day +100. Plerixafor-based mobilization is associated with slightly reduced number of CFU-GM and minimal delay in engraftment that is independent of CD34+ cell dose. Hematopoietic graft function on Day 100 is not affected by mobilization strategy. © 2011 American Association of Blood Banks.

Red Blood Cell Function and Dysfunction: Redox Regulation, Nitric Oxide Metabolism, Anemia

PubMed Central

Kuhn, Viktoria; Diederich, Lukas; Keller, T.C. Stevenson; Kramer, Christian M.; Lückstädt, Wiebke; Panknin, Christina; Suvorava, Tatsiana; Isakson, Brant E.; Kelm, Malte

2017-01-01

Abstract Significance: Recent clinical evidence identified anemia to be correlated with severe complications of cardiovascular disease (CVD) such as bleeding, thromboembolic events, stroke, hypertension, arrhythmias, and inflammation, particularly in elderly patients. The underlying mechanisms of these complications are largely unidentified. Recent Advances: Previously, red blood cells (RBCs) were considered exclusively as transporters of oxygen and nutrients to the tissues. More recent experimental evidence indicates that RBCs are important interorgan communication systems with additional functions, including participation in control of systemic nitric oxide metabolism, redox regulation, blood rheology, and viscosity. In this article, we aim to revise and discuss the potential impact of these noncanonical functions of RBCs and their dysfunction in the cardiovascular system and in anemia. Critical Issues: The mechanistic links between changes of RBC functional properties and cardiovascular complications related to anemia have not been untangled so far. Future Directions: To allow a better understanding of the complications associated with anemia in CVD, basic and translational science studies should be focused on identifying the role of noncanonical functions of RBCs in the cardiovascular system and on defining intrinsic and/or systemic dysfunction of RBCs in anemia and its relationship to CVD both in animal models and clinical settings. Antioxid. Redox Signal. 26, 718–742. PMID:27889956

IMPROVED BONDING METHOD

DOEpatents

Padgett, E.V. Jr.; Warf, D.H.

1964-04-28

An improved process of bonding aluminum to aluminum without fusion by ultrasonic vibrations plus pressure is described. The surfaces to be bonded are coated with an aqueous solution of alkali metal stearate prior to assembling for bonding. (AEC) O H19504 Present information is reviewed on steady state proliferation, differentiation, and maturation of blood cells in mammals. Data are cited from metabolic tracer studies, autoradiographic studies, cytologic studies, studies of hematopoietic response to radiation injuries, and computer analyses of blood cell production. A 3-step model for erythropoiesis and a model for granulocyte kinetics are presented. New approaches to the study of lymphocytopoiesis described include extracorporeal blood irradiation to deplete lymphocytic tissue without direct injury to the formative tissues as a means to study the stressed system, function control, and rates of proliferation. It is pointed out that present knowledge indicates that lymphocytes comprise a mixed family, with diverse life spans, functions, and migration patterns with apparent aimless recycling from modes to lymph to blood to nodes that has not yet been quantitated. Areas of future research are postulated. (70 references.) (C.H.)

Effects of dietary almond- and olive oil-based docosahexaenoic acid- and vitamin E-enriched beverage supplementation on athletic performance and oxidative stress markers.

PubMed

Capó, X; Martorell, M; Busquets-Cortés, C; Sureda, A; Riera, J; Drobnic, F; Tur, J A; Pons, A

2016-12-07

Functional beverages based on almonds and olive oil and enriched with α-tocopherol and docosahexaenoic acid (DHA) could be useful in modulating oxidative stress and enhancing physical performance in sportsmen. The aim of this work was to evaluate the effects of supplementation with functional beverages on physical performance, plasma and erythrocyte fatty acids' and polyphenol handling, oxidative and nitrative damage, and antioxidant and mitochondrial gene expression in young and senior athletes. Athletes performed maximal exercise tests before and after one month of dietary supplementation and blood samples were taken immediately before and one hour after each test. The beverages did not alter performance parameters during maximal exercise. Supplementation increased polyunsaturated and reduced saturated plasma fatty acids while increasing the DHA erythrocyte content; it maintained basal plasma and blood polyphenol levels, but increased the blood cell polyphenol concentration in senior athletes. Supplementation protects against oxidative damage although it enhances nitrative damage in young athletes. The beverages enhance the gene expression of antioxidant enzymes in peripheral blood mononuclear cells after exercise in young athletes.

A pilot study of dose-intensified procarbazine, CCNU, vincristine for poor prognosis brain tumors utilizing fibronectin-assisted, retroviral-mediated modification of CD34+ peripheral blood cells with O6-methylguanine DNA methyltransferase.

PubMed

Cornetta, K; Croop, J; Dropcho, E; Abonour, R; Kieran, M W; Kreissman, S; Reeves, L; Erickson, L C; Williams, D A

2006-09-01

Administration of chemotherapy is often limited by myelosuppression. Expression of drug-resistance genes in hematopoietic cells has been proposed as a means to decrease the toxicity of cytotoxic agents. In this pilot study, we utilized a retroviral vector expressing methylguanine DNA methyltransferase (MGMT) to transduce hematopoietic progenitors, which were subsequently used in the setting of alkylator therapy (procarbazine, CCNU, vincristine (PCV)) for poor prognosis brain tumors. Granulocyte colony-stimulating factor (G-CSF)-mobilized peripheral blood progenitor cells were collected by apheresis and enriched for CD34+ expression. Nine subjects were infused with CD34+-enriched cells treated in a transduction procedure involving a 4-day exposure to cytokines with vector exposure on days 3 and 4. No major adverse event was related to the gene therapy procedure. Importantly, the engraftment kinetics of the treated product was similar to unmanipulated peripheral blood stem cells, suggesting that the ex vivo manipulation did not significantly reduce engrafting progenitor cell function. Gene-transduced cells were detected in all subjects. Although the level and duration was limited, patients receiving cells transduced using fibronectin 'preloaded' with virus supernatant appeared to show improved in vivo marking frequency. These findings demonstrate the feasibility and safety of utilizing MGMT-transduced CD34+ peripheral blood progenitor cells in the setting of chemotherapy.

Control of RUNX-induced repression of Notch signaling by MLF and its partner DnaJ-1 during Drosophila hematopoiesis

PubMed Central

Gobert, Vanessa; Augé, Benoit; Burlet-Schiltz, Odile; Haenlin, Marc

2017-01-01

A tight regulation of transcription factor activity is critical for proper development. For instance, modifications of RUNX transcription factors dosage are associated with several diseases, including hematopoietic malignancies. In Drosophila, Myeloid Leukemia Factor (MLF) has been shown to control blood cell development by stabilizing the RUNX transcription factor Lozenge (Lz). However, the mechanism of action of this conserved family of proteins involved in leukemia remains largely unknown. Here we further characterized MLF’s mode of action in Drosophila blood cells using proteomic, transcriptomic and genetic approaches. Our results show that MLF and the Hsp40 co-chaperone family member DnaJ-1 interact through conserved domains and we demonstrate that both proteins bind and stabilize Lz in cell culture, suggesting that MLF and DnaJ-1 form a chaperone complex that directly regulates Lz activity. Importantly, dnaj-1 loss causes an increase in Lz+ blood cell number and size similarly as in mlf mutant larvae. Moreover we find that dnaj-1 genetically interacts with mlf to control Lz level and Lz+ blood cell development in vivo. In addition, we show that mlf and dnaj-1 loss alters Lz+ cell differentiation and that the increase in Lz+ blood cell number and size observed in these mutants is caused by an overactivation of the Notch signaling pathway. Finally, using different conditions to manipulate Lz activity, we show that high levels of Lz are required to repress Notch transcription and signaling. All together, our data indicate that the MLF/DnaJ-1-dependent increase in Lz level allows the repression of Notch expression and signaling to prevent aberrant blood cell development. Thus our findings establish a functional link between MLF and the co-chaperone DnaJ-1 to control RUNX transcription factor activity and Notch signaling during blood cell development in vivo. PMID:28742844

Mechanical properties of stored red blood cells using optical tweezers

NASA Astrophysics Data System (ADS)

Fontes, Adriana; Alexandre de Thomaz, Andre; de Ysasa Pozzo, Liliana; de Lourdes Barjas-Castro, Maria; Brandao, Marcelo M.; Saad, Sara T. O.; Barbosa, Luiz Carlos; Cesar, Carlos Lenz

2005-08-01

We have developed a method for measuring the red blood cell (RBC) membrane overall elasticity μ by measuring the deformation of the cells when dragged at a constant velocity through a plasma fluid by an optical tweezers. The deformability of erythrocytes is a critical determinant of blood flow in the microcirculation. We tested our method and hydrodynamic models, which included the presence of two walls, by measuring the RBC deformation as a function of drag velocity and of the distance to the walls. The capability and sensitivity of this method can be evaluated by its application to a variety of studies, such as, the measurement of RBC elasticity of sickle cell anemia patients comparing homozygous (HbSS), including patients taking hydroxyrea (HU) and heterozygous (HbAS) with normal donors and the RBC elasticity measurement of gamma irradiated stored blood for transfusion to immunosupressed patients as a function of time and dose. These studies show that the technique has the sensitivity to discriminate heterozygous and homozygous sickle cell anemia patients from normal donors and even follow the course of HU treatment of Homozygous patients. The gamma irradiation studies show that there is no significant change in RBC elasticity over time for up to 14 days of storage, regardless of whether the unit was irradiated or not, but there was a huge change in the measured elasticity for the RBC units stored for more than 21 days after irradiation. These finds are important for the assessment of stored irradiated RBC viability for transfusion purposes because the present protocol consider 28 storage days after irradiation as the limit for the RBC usage.

l-Arginine-Dependent Epigenetic Regulation of Interleukin-10, but Not Transforming Growth Factor-β, Production by Neonatal Regulatory T Lymphocytes

PubMed Central

Yu, Hong-Ren; Tsai, Ching-Chang; Chang, Ling-Sai; Huang, Hsin-Chun; Cheng, Hsin-Hsin; Wang, Jiu-Yao; Sheen, Jiunn-Ming; Kuo, Ho-Chang; Hsieh, Kai-Sheng; Huang, Ying-Hsien; Yang, Kuender D.; Hsu, Te-Yao

2017-01-01

A growing number of diseases in humans, including trauma, certain cancers, and infection, are known to be associated with l-arginine deficiency. In addition, l-arginine must be supplemented by diet during pregnancy to aid fetal development. In conditions of l-arginine depletion, T cell proliferation is impaired. We have previously shown that neonatal blood has lower l-arginine levels than adult blood, which is associated with poor neonatal lymphocyte proliferation, and that l-arginine enhances neonatal lymphocyte proliferation through an interleukin (IL)-2-independent pathway. In this study, we have further investigated how exogenous l-arginine enhances neonatal regulatory T-cells (Tregs) function in relation to IL-10 production under epigenetic regulation. Results showed that cord blood mononuclear cells (CBMCs) produced higher levels of IL-10 than adult peripheral blood mononuclear cells (PBMCs) by phytohemagglutinin stimulation but not by anti-CD3/anti-CD28 stimulation. Addition of exogenous l-arginine had no effect on transforming growth factor-β production by PBMCs or CBMCs, but enhanced IL-10 production by neonatal CD4+CD25+FoxP3+ Tregs. Further studies showed that IL-10 promoter DNA hypomethylation, rather than histone modification, corresponded to the l-arginine-induced increase in IL-10 production by neonatal CD4+ T cells. These results suggest that l-arginine modulates neonatal Tregs through the regulation of IL-10 promoter DNA methylation. l-arginine supplementation may correct the Treg function in newborns with l-arginine deficiency. PMID:28487700

In vitro models of the blood–brain barrier: An overview of commonly used brain endothelial cell culture models and guidelines for their use

PubMed Central

Helms, Hans C; Abbott, N Joan; Burek, Malgorzata; Cecchelli, Romeo; Couraud, Pierre-Olivier; Deli, Maria A; Förster, Carola; Galla, Hans J; Romero, Ignacio A; Shusta, Eric V; Stebbins, Matthew J; Vandenhaute, Elodie; Weksler, Babette

2016-01-01

The endothelial cells lining the brain capillaries separate the blood from the brain parenchyma. The endothelial monolayer of the brain capillaries serves both as a crucial interface for exchange of nutrients, gases, and metabolites between blood and brain, and as a barrier for neurotoxic components of plasma and xenobiotics. This “blood-brain barrier” function is a major hindrance for drug uptake into the brain parenchyma. Cell culture models, based on either primary cells or immortalized brain endothelial cell lines, have been developed, in order to facilitate in vitro studies of drug transport to the brain and studies of endothelial cell biology and pathophysiology. In this review, we aim to give an overview of established in vitro blood–brain barrier models with a focus on their validation regarding a set of well-established blood–brain barrier characteristics. As an ideal cell culture model of the blood–brain barrier is yet to be developed, we also aim to give an overview of the advantages and drawbacks of the different models described. PMID:26868179

THE SKIN | Functional morphology of the integumentary system in fishes

USGS Publications Warehouse

Elliott, D.G.; Farrell, Anthony P.

2011-01-01

The integument that covers the outer surface of a fish’s body and fins is a multifunctional organ, with morphological features highly adapted to carry out these functions. The integument consists of two layers. The outer layer, the epidermis, is essentially cellular in structure, comprised of a multilayered epithelium that usually includes specialized cells. The inner layer, the dermis, is primarily a fibrous structure with relatively few cells, although it may contain scales, nerves, blood vessels, adipose tissue, and pigment cells.

[Blood-nerve barrier: structure and function].

PubMed

Kanda, Takashi

2011-06-01

The blood-nerve barrier (BNB) is a dynamic interface between the endoneurial microenvironment and surrounding extracellular space or blood contents, and is localized the innermost layer of multilayered ensheathing perineurium and endoneurial microvessels. Since the BNB is a key structure controlling the internal milieu of the peripheral nerve parenchyma, adequate understanding of the BNB is crucial for developing treatment strategies for human peripheral nervous system disorders, including Guillain-Barré syndrome, chronic inflammatory demyelinating polyneuropathy, and diabetic and various metabolic/toxic neuropathies. However, fewer studies have been conducted on the BNB, if we compare against the number of studies on the blood-brain barrier. This is because of the lack of adequate human cell lines originating from the BNB. In our laboratory, human immortal cell lines from the BNB, namely, the endothelial cell line and pericyte cell line, have recently been established and vigorous investigations of their biological and physiological properties are now underway. Pericytes constituting the BNB were found to possess robust ability of controlling BNB integrity via secretion of various cytokines and growth factors including bFGF, VEGF, GDNF, BDNF, and angiopoietin-1. Unknown soluble factors secreted by pericytes also contribute to the upregulation of claudin-5 in endothelial cells in the BNB and thus, strengthen the barrier function of the BNB. In diabetic neuropathy, pericytes were shown to regulate the vascular basement membrane, while AGEs were shown to induce basement membrane hypertrophy and disrupt the BNB by increasing the autocrine secretion of VEGF and TGF-beta from pericytes. In this review article, we discuss the macroscopic and microscopic anatomy of the human BNB as well as the molecular mechanisms of mononuclear cell infiltration across the BNB.

Blood flow and microgravity

NASA Astrophysics Data System (ADS)

Bureau, Lionel; Coupier, Gwennou; Dubois, Frank; Duperray, Alain; Farutin, Alexander; Minetti, Christophe; Misbah, Chaouqi; Podgorski, Thomas; Tsvirkun, Daria; Vysokikh, Mikhail

2017-01-01

The absence of gravity during space flight can alter cardio-vascular functions partially due to reduced physical activity. This affects the overall hemodynamics, and in particular the level of shear stresses to which blood vessels are submitted. Long-term exposure to space environment is thus susceptible to induce vascular remodeling through a mechanotransduction cascade that couples vessel shape and function with the mechanical cues exerted by the circulating cells on the vessel walls. Central to such processes, the glycocalyx - i.e. the micron-thick layer of biomacromolecules that lines the lumen of blood vessels and is directly exposed to blood flow - is a major actor in the regulation of biochemical and mechanical interactions. We discuss in this article several experiments performed under microgravity, such as the determination of lift force and collective motion in blood flow, and some preliminary results obtained in artificial microfluidic circuits functionalized with endothelium that offer interesting perspectives for the study of the interactions between blood and endothelium in healthy condition as well as by mimicking the degradation of glycocalyx caused by long space missions. A direct comparison between experiments and simulations is discussed. xml:lang="fr"

Epstein-Barr Virus-positive T-cell Lymphoproliferative Disease Following Umbilical Cord Blood Transplantation for Acute Myeloid Leukemia.

PubMed

Yui, Shunsuke; Yamaguchi, Hiroki; Imadome, Ken-ichi; Arai, Ayako; Takahashi, Mikiko; Ohashi, Ryuji; Tamai, Hayato; Moriya, Keiichi; Nakayama, Kazutaka; Shimizu, Akira; Inokuchi, Koiti

2016-01-01

We report a case of the extremely rare condition Epstein-Barr virus (EBV)-positive T-cell lymphoproliferative disease (LPD) which occurred after umbilical cord blood transplantation. A 25-year-old Japanese man underwent cord blood transplantation from a male human leukocyte antigen 4/6-matched donor due to acute myeloid leukemia with trisomy 8. Bone marrow examination on day 30 showed chimerism with at least 90% donor cells and complete hematological response. Chronic symptoms of graft-versus-host disease appeared only on the skin and were successfully treated with cyclosporine alone. Three years later, however, the patient experienced repeated cold-like symptoms and was hospitalized with liver dysfunction. A high fever developed and was followed by significant edema of the right side of the face. The EBV DNA copy number in whole peripheral blood was 2×10(4)/mL. Liver biopsy showed invasion of EBV-infected CD8-positive T cells. Southern blotting analysis of the whole peripheral blood showed that the T-cell receptor Cβ1 rearrangement was positive. On the basis of these results, EBV-positive T-cell LPD was diagnosed and treated with prednisolone, cyclosporine, and etoposide, followed by cyclophosphamide, doxorubicin, vincristine, and prednisone. However, the patient died of cardiac function failure, pneumonia, and pulmonary hemorrhage, all of unidentified cause. Most cases of EBV-related LPD after hematopoietic stem cell transplantation consist of EBV-positive B-cell LPD, and, to our knowledge, de novo EBV-positive T-cell LPD subsequent to transplantation has not been previously reported.

Biologic properties of endothelial progenitor cells and their potential for cell therapy.

PubMed

Young, Pampee P; Vaughan, Douglas E; Hatzopoulos, Antonis K

2007-01-01

Recent studies indicate that portions of ischemic and tumor neovasculature are derived by neovasculogenesis, whereby bone marrow (BM)-derived circulating endothelial progenitor cells (EPCs) home to sites of regenerative or malignant growth and contribute to blood vessel formation. Recent data from animal models suggest that a variety of cell types, including unfractionated BM mononuclear cells and those obtained by ex vivo expansion of human peripheral blood or enriched progenitors, can function as EPCs to promote tissue vasculogenesis, regeneration, and repair when introduced in vivo. The promising preclinical results have led to several human clinical trials using BM as a potential source of EPCs in cardiac repair as well as ongoing basic research on using EPCs in tissue engineering or as cell therapy to target tumor growth.

Glioblastoma: A Pathogenic Crosstalk between Tumor Cells and Pericytes

PubMed Central

Redondo-Garcia, Carolina; Martinez, Salvador

2014-01-01

Cancers likely originate in progenitor zones containing stem cells and perivascular stromal cells. Much evidence suggests stromal cells play a central role in tumor initiation and progression. Brain perivascular cells (pericytes) are contractile and function normally to regulate vessel tone and morphology, have stem cell properties, are interconvertible with macrophages and are involved in new vessel formation during angiogenesis. Nevertheless, how pericytes contribute to brain tumor infiltration is not known. In this study we have investigated the underlying mechanism by which the most lethal brain cancer, Glioblastoma Multiforme (GBM) interacts with pre-existing blood vessels (co-option) to promote tumor initiation and progression. Here, using mouse xenografts and laminin-coated silicone substrates, we show that GBM malignancy proceeds via specific and previously unknown interactions of tumor cells with brain pericytes. Two-photon and confocal live imaging revealed that GBM cells employ novel, Cdc42-dependent and actin-based cytoplasmic extensions, that we call flectopodia, to modify the normal contractile activity of pericytes. This results in the co-option of modified pre-existing blood vessels that support the expansion of the tumor margin. Furthermore, our data provide evidence for GBM cell/pericyte fusion-hybrids, some of which are located on abnormally constricted vessels ahead of the tumor and linked to tumor-promoting hypoxia. Remarkably, inhibiting Cdc42 function impairs vessel co-option and converts pericytes to a phagocytic/macrophage-like phenotype, thus favoring an innate immune response against the tumor. Our work, therefore, identifies for the first time a key GBM contact-dependent interaction that switches pericyte function from tumor-suppressor to tumor-promoter, indicating that GBM may harbor the seeds of its own destruction. These data support the development of therapeutic strategies directed against co-option (preventing incorporation and modification of pre-existing blood vessels), possibly in combination with anti-angiogenesis (blocking new vessel formation), which could lead to improved vascular targeting not only in Glioblastoma but also for other cancers. PMID:25032689

Pressure-driven occlusive flow of a confined red blood cell.

PubMed

Savin, Thierry; Bandi, M M; Mahadevan, L

2016-01-14

When red blood cells (RBCs) move through narrow capillaries in the microcirculation, they deform as they flow. In pathophysiological processes such as sickle cell disease and malaria, RBC motion and flow are severely restricted. To understand this threshold of occlusion, we use a combination of experiment and theory to study the motion of a single swollen RBC through a narrow glass capillary of varying inner diameter. By tracking the movement of the squeezed cell as it is driven by a controlled pressure drop, we measure the RBC velocity as a function of the pressure gradient as well as the local capillary diameter, and find that the effective blood viscosity in this regime increases with both decreasing RBC velocity and tube radius by following a power-law that depends upon the length of the confined cell. Our observations are consistent with a simple elasto-hydrodynamic model and highlight the role of lateral confinement in the occluded pressure-driven slow flow of soft confined objects.

Drosophila sessile hemocyte clusters are true hematopoietic tissues that regulate larval blood cell differentiation

PubMed Central

Leitão, Alexandre B; Sucena, Élio

2015-01-01

Virtually all species of coelomate animals contain blood cells that display a division of labor necessary for homeostasis. This functional partition depends upon the balance between proliferation and differentiation mostly accomplished in the hematopoietic organs. In Drosophila melanogaster, the lymph gland produces plasmatocytes and crystal cells that are not released until pupariation. Yet, throughout larval development, both hemocyte types increase in numbers. Mature plasmatocytes can proliferate but it is not known if crystal cell numbers increase by self-renewal or by de novo differentiation. We show that new crystal cells in third instar larvae originate through a Notch-dependent process of plasmatocyte transdifferentiation. This process occurs in the sessile clusters and is contingent upon the integrity of these structures. The existence of this hematopoietic tissue, relying on structure-dependent signaling events to promote blood homeostasis, creates a new paradigm for addressing outstanding questions in Drosophila hematopoiesis and establishing further parallels with vertebrate systems. DOI: http://dx.doi.org/10.7554/eLife.06166.001 PMID:25650737

[Comparative ultrastructural study of parotid gland, lacrimal gland and pituitary gland between miniature pig and mouse].

PubMed

Yan, Xing; Hai, Bo; Sun, Yi-lin; Zhang, Chun-mei; Wang, Song-ling

2009-02-01

To study the ultrastructure of parotid glands, lacrimal glands and pituitary glands between miniature pig and mouse. Five adult miniature pigs and 5 mice were studied. Ultrastructure of their parotid glands, lacrimal glands, and pituitary glands was observed. The secretary granules in acinar cell of miniature pig parotid glands showed higher density and more aequalis than those of mice. The cell apparatus in acinar cell of mouse parotid glands were more plentiful than those of miniature pigs. The secretary granules on blood vessel wall were richer in parotid gland of miniature pigs compared with mouse parotid gland. Lacrimal gland had the similar ultrastructure to parotid gland in these two animals. Many blood vessel antrum were found in pituitary glands of these two animals. Compared with mouse parotid glands, there are more secretary granules in acinar cells and vascular endothelial cells in miniature pig parotid glands, which might enter blood stream and have function of endocrine secretion.

Cell–specific Variation in E-selectin Ligand Expression Among Human Peripheral Blood Mononuclear Cells: Implications for Immunosurveillance and Pathobiology

PubMed Central

Silva, Mariana; Fung, Ronald Kam Fai; Donnelly, Conor Brian; Videira, Paula Alexandra; Sackstein, Robert

2017-01-01

Both host defense and immunopathology are shaped by the ordered recruitment of circulating leukocytes to affected sites, a process initiated by binding of blood-borne cells to E-selectin displayed at target endothelial beds. Accordingly, knowledge of the expression and function of leukocyte E-selectin ligands is key to understanding the tempo and specificity of immunoreactivity. Here, we performed E-selectin adherence assays under hemodynamic flow conditions coupled with flow cytometry and western blot analysis to elucidate the function and structural biology of glycoprotein E-selectin ligands expressed on human peripheral blood mononuclear cells (PBMCs). Circulating monocytes uniformly express high levels of the canonical E-selectin binding determinant sLeX and display markedly greater adhesive interactions with E-selectin than do circulating lymphocytes, which exhibit variable E-selectin binding among CD4+ and CD8+ T-cells but no binding by B-cells. Monocytes prominently present sLeX decorations on an array of protein scaffolds including PSGL-1, CD43, and CD44 (rendering the E-selectin ligands CLA, CD43E, and HCELL, respectively), and B-cells altogether lack E-selectin ligands. Quantitative PCR gene expression studies of glycosyltransferases that regulate display of sLeX reveal high transcript levels among circulating monocytes and low levels among circulating B-cells, and, commensurately, cell surface α(1,3)-fucosylation reveals that acceptor sialyllactosaminyl glycans convertible into sLeX are abundantly expressed on human monocytes yet are relatively deficient on B-cells. Collectively, these findings unveil distinct cell-specific patterns of E-selectin ligand expression among human PBMCs, indicating that circulating monocytes are specialized to engage E-selectin and providing key insights into the molecular effectors mediating recruitment of these cells at inflammatory sites. PMID:28330896

Donor-specific hypo-responsiveness occurs in simultaneous liver-kidney transplant recipients after the first year.

PubMed

Taner, Timucin; Gustafson, Michael P; Hansen, Michael J; Park, Walter D; Bornschlegl, Svetlana; Dietz, Allan B; Stegall, Mark D

2018-06-01

Kidney allografts of patients who undergo simultaneous liver-kidney transplantation incur less immune-mediated injury, and retain better function compared to other kidney allografts. To characterize the host alloimmune responses in 28 of these patients, we measured the donor-specific alloresponsiveness and phenotypes of peripheral blood cells after the first year. These values were then compared to those of 61 similarly immunosuppressed recipients of a solitary kidney or 31 recipients of liver allografts. Four multicolor, non-overlapping flow cytometry protocols were used to assess the immunophenotypes. Mixed cell cultures with donor or third party cells were used to measure cell proliferation and interferon gamma production. Despite a significant overlap, simultaneous liver-kidney transplant recipients had a lower overall frequency of circulating CD8 + , activated CD4 + and effector memory T cells, compared to solitary kidney transplant recipients. Simultaneous liver-kidney transplant recipient T cells had a significantly lower proliferative response to the donor cells compared to solitary kidney recipients (11.9 vs. 42.9%), although their response to third party cells was unaltered. The frequency of interferon gamma producing alloreactive T cells in simultaneous liver-kidney transplant recipients was significantly lower than that of solitary kidney transplant recipients. Flow cytometric analysis of the mixed cultures demonstrated that both alloreactive CD4 + and CD8 + compartments of the simultaneous liver-kidney transplant recipient circulating blood cells were smaller. Thus, the phenotypic and functional characteristics of the circulating blood cells of the simultaneous liver-kidney transplant recipients resembled those of solitary liver transplant recipients, and appear to be associated with donor-specific hypo-alloresponsiveness. Copyright © 2018 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Enrichment isolation of adipose-derived stem/stromal cells from the liquid portion of liposuction aspirates with the use of an adherent column.

PubMed

Doi, Kentaro; Kuno, Shinichiro; Kobayashi, Akira; Hamabuchi, Takahisa; Kato, Harunosuke; Kinoshita, Kahori; Eto, Hitomi; Aoi, Noriyuki; Yoshimura, Kotaro

2014-03-01

Adipose-derived stem/progenitor cells (ASCs) are typically obtained from the lipoaspirates; however, a smaller number of ASCs can be isolated without enzymatic digestion from the infranatant liposuction aspirate fluid (LAF). We evaluated the effectiveness of an adherent column, currently used to isolate mesenchymal stromal cells from bone marrow, to isolate LAF cells. We applied peripheral blood (PB), PB mixed with cultured ASCs (PB-ASC), and LAF solution to the column and divided it into two fractions, the adherent (positive) and the non-adherent (negative) fractions. We compared this method with hypotonic hemolysis (lysis) for the red blood cell count, nucleated cells count and cell compositions as well as functional properties of isolated mesenchymal cells. The column effectively removed red blood cells, though the removal efficiency was slightly inferior to hemolysis. After column processing of PB-ASC, 60.5% of ASCs (53.2% by lysis) were selectively collected in the positive fraction, and the negative fraction contained almost no ASCs. After processing of LAF solution, nucleated cell yields were comparable between the column and hemolysis; however, subsequent adherent culture indicated that a higher average ASC yield was obtained from the column-positive samples than from the lysis samples, suggesting that the column method may be superior to hemolysis for obtaining viable ASCs. Mesenchymal differentiation and network formation assays showed no statistical differences in ASC functions between the lysis and column-positive samples. Our results suggest that a column with non-woven rayon and polyethylene fabrics is useful for isolating stromal vascular fraction cells from LAF solutions for clinical applications. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

High expression of NKG2A/CD94 and low expression of granzyme B are associated with reduced cord blood NK cell activity.

PubMed

Wang, Yanyan; Xu, Han; Zheng, Xiaodong; Wei, Haiming; Sun, Rui; Tian, Zhigang

2007-10-01

Human umbilical cord blood (CB) has recently been used as a source of stem cells in transplantation. NK cells derived from CB are the key effector cells involved in graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL). It was reported that the activity of CB NK cells was lower than that of adult peripheral blood (PB) NK cells. In this study, we analyzed the expression of some NK cell receptors and cytotoxicity-related molecules in CB and PB NK cells. The expressions of activating NK receptors, CD16, NKG2D and NKp46, did not show significant difference between CB and PB NK cells. But the expression of inhibitory receptor NKG2A/CD94 was significantly higher on CB NK cells. As to the effector function molecules, granzyme B was expressed significantly lower in CB NK cells, but the expressions of intracellular perforin, IFN-gamma, TNF-alpha and cell surface FasL and TRAIL did not show difference between CB and PB NK cells. The results indicated that the high expression of NKG2A/CD94 and low expression of granzyme B may be related with the reduced activity of CB NK cells.

Transcriptional profiling of CD31(+) cells isolated from murine embryonic stem cells.

PubMed

Mariappan, Devi; Winkler, Johannes; Chen, Shuhua; Schulz, Herbert; Hescheler, Jürgen; Sachinidis, Agapios

2009-02-01

Identification of genes involved in endothelial differentiation is of great interest for the understanding of the cellular and molecular mechanisms involved in the development of new blood vessels. Mouse embryonic stem (mES) cells serve as a potential source of endothelial cells for transcriptomic analysis. We isolated endothelial cells from 8-days old embryoid bodies by immuno-magnetic separation using platelet endothelial cell adhesion molecule-1 (also known as CD31) expressed on both early and mature endothelial cells. CD31(+) cells exhibit endothelial-like behavior by being able to incorporate DiI-labeled acetylated low-density lipoprotein as well as form tubular structures on matrigel. Quantitative and semi-quantitative PCR analysis further demonstrated the increased expression of endothelial transcripts. To ascertain the specific transcriptomic identity of the CD31(+) cells, large-scale microarray analysis was carried out. Comparative bioinformatic analysis reveals an enrichment of the gene ontology categories angiogenesis, blood vessel morphogenesis, vasculogenesis and blood coagulation in the CD31(+) cell population. Based on the transcriptomic signatures of the CD31(+) cells, we conclude that this ES cell-derived population contains endothelial-like cells expressing a mesodermal marker BMP2 and possess an angiogenic potential. The transcriptomic characterization of CD31(+) cells enables an in vitro functional genomic model to identify genes required for angiogenesis.

Transplantation of autologous bone marrow-derived mesenchymal stem cells for traumatic brain injury☆

PubMed Central

Jiang, Jindou; Bu, Xingyao; Liu, Meng; Cheng, Peixun

2012-01-01

Results from the present study demonstrated that transplantation of autologous bone marrow-derived mesenchymal stem cells into the lesion site in rat brain significantly ameliorated brain tissue pathological changes and brain edema, attenuated glial cell proliferation, and increased brain-derived neurotrophic factor expression. In addition, the number of cells double-labeled for 5-bromodeoxyuridine/glial fibrillary acidic protein and cells expressing nestin increased. Finally, blood vessels were newly generated, and the rats exhibited improved motor and cognitive functions. These results suggested that transplantation of autologous bone marrow-derived mesenchymal stem cells promoted brain remodeling and improved neurological functions following traumatic brain injury. PMID:25806058

Endothelial dysfunction and amyloid-β-induced neurovascular alterations

PubMed Central

Koizumi, Kenzo; Wang, Gang; Park, Laibaik

2015-01-01

Alzheimer's disease (AD) and cerebrovascular diseases share common vascular risk factors that have disastrous effects on cerebrovascular regulation. Endothelial cells, lining inner walls of cerebral blood vessels, form a dynamic interface between the blood and the brain and are critical for the maintenance of neurovascular homeostasis. Accordingly, injury in endothelial cells is regarded as one of the earliest symptoms of impaired vasoregulatory mechanisms. Extracellular buildup of amyloid-β (Aβ) is a central pathogenic factor in AD. Aβ exerts potent detrimental effects on cerebral blood vessels and impairs endothelial structure and function. Recent evidence implicates vascular oxidative stress and activation of the nonselective cationic channel transient receptor potential melastatin (TRPM)-2 on endothelial cells in the mechanisms of Aβ-induced neurovascular dysfunction. Thus, Aβ triggers opening of TRPM2 channels in endothelial cells leading to intracellular Ca2+ overload and vasomotor dysfunction. The cerebrovascular dysfunction may contribute to AD pathogenesis by reducing the cerebral blood supply, leading to increased susceptibility to vascular insufficiency, and by promoting Aβ accumulation. The recent realization that vascular factors contribute to AD pathobiology suggests new targets for the prevention and treatment of this devastating disease. PMID:26328781

The non-coding RNA landscape of human hematopoiesis and leukemia.

PubMed

Schwarzer, Adrian; Emmrich, Stephan; Schmidt, Franziska; Beck, Dominik; Ng, Michelle; Reimer, Christina; Adams, Felix Ferdinand; Grasedieck, Sarah; Witte, Damian; Käbler, Sebastian; Wong, Jason W H; Shah, Anushi; Huang, Yizhou; Jammal, Razan; Maroz, Aliaksandra; Jongen-Lavrencic, Mojca; Schambach, Axel; Kuchenbauer, Florian; Pimanda, John E; Reinhardt, Dirk; Heckl, Dirk; Klusmann, Jan-Henning

2017-08-09

Non-coding RNAs have emerged as crucial regulators of gene expression and cell fate decisions. However, their expression patterns and regulatory functions during normal and malignant human hematopoiesis are incompletely understood. Here we present a comprehensive resource defining the non-coding RNA landscape of the human hematopoietic system. Based on highly specific non-coding RNA expression portraits per blood cell population, we identify unique fingerprint non-coding RNAs-such as LINC00173 in granulocytes-and assign these to critical regulatory circuits involved in blood homeostasis. Following the incorporation of acute myeloid leukemia samples into the landscape, we further uncover prognostically relevant non-coding RNA stem cell signatures shared between acute myeloid leukemia blasts and healthy hematopoietic stem cells. Our findings highlight the importance of the non-coding transcriptome in the formation and maintenance of the human blood hierarchy.While micro-RNAs are known regulators of haematopoiesis and leukemogenesis, the role of long non-coding RNAs is less clear. Here the authors provide a non-coding RNA expression landscape of the human hematopoietic system, highlighting their role in the formation and maintenance of the human blood hierarchy.

Isolation and characterization of naturally occurring subclasses of human peripheral blood T cells with regulatory functions.

PubMed

Strelkauskas, A J; Schauf, V; Wilson, B S; Chess, L; Schlossman, S F

1978-04-01

By utilizing naturally occurring autoimmune antibodies from patients with juvenile rheumatoid arthritis, we have isolated and functionally characterized two unique subpopulations of T cells. JRA+ T cells, i.e., those identified by sera from these patients, react poorly in response to allogeneic cells, respond to Con A but not PHA, and do not help in the synthesis and secretion of Ig by B cells. In contrast, JRA- T cells, i.e., those not identified by sera from these patients, respond very well to allogeneic cells, proliferate well in response to PHA but not Con A, and more interestingly, can greatly enhance the secretion of Ig by B cells.

Transcriptomic meta-analysis identifies gene expression characteristics in various samples of HIV-infected patients with nonprogressive disease.

PubMed

Zhang, Le-Le; Zhang, Zi-Ning; Wu, Xian; Jiang, Yong-Jun; Fu, Ya-Jing; Shang, Hong

2017-09-12

A small proportion of HIV-infected patients remain clinically and/or immunologically stable for years, including elite controllers (ECs) who have undetectable viremia (<50 copies/ml) and long-term nonprogressors (LTNPs) who maintain normal CD4 + T cell counts for prolonged periods (>10 years). However, the mechanism of nonprogression needs to be further resolved. In this study, a transcriptome meta-analysis was performed on nonprogressor and progressor microarray data to identify differential transcriptome pathways and potential biomarkers. Using the INMEX (integrative meta-analysis of expression data) program, we performed the meta-analysis to identify consistently differentially expressed genes (DEGs) in nonprogressors and further performed functional interpretation (gene ontology analysis and pathway analysis) of the DEGs identified in the meta-analysis. Five microarray datasets (81 cases and 98 controls in total), including whole blood, CD4 + and CD8 + T cells, were collected for meta-analysis. We determined that nonprogressors have reduced expression of important interferon-stimulated genes (ISGs), CD38, lymphocyte activation gene 3 (LAG-3) in whole blood, CD4 + and CD8 + T cells. Gene ontology (GO) analysis showed a significant enrichment in DEGs that function in the type I interferon signaling pathway. Upregulated pathways, including the PI3K-Akt signaling pathway in whole blood, cytokine-cytokine receptor interaction in CD4 + T cells and the MAPK signaling pathway in CD8 + T cells, were identified in nonprogressors compared with progressors. In each metabolic functional category, the number of downregulated DEGs was more than the upregulated DEGs, and almost all genes were downregulated DEGs in the oxidative phosphorylation (OXPHOS) and tricarboxylic acid (TCA) cycle in the three types of samples. Our transcriptomic meta-analysis provides a comprehensive evaluation of the gene expression profiles in major blood types of nonprogressors, providing new insights in the understanding of HIV pathogenesis and developing strategies to delay HIV disease progression.

Decreased level of cord blood circulating endothelial colony-forming cells in preeclampsia.

PubMed

Muñoz-Hernandez, Rocio; Miranda, Maria L; Stiefel, Pablo; Lin, Ruei-Zeng; Praena-Fernández, Juan M; Dominguez-Simeon, Maria J; Villar, Jose; Moreno-Luna, Rafael; Melero-Martin, Juan M

2014-07-01

Preeclampsia is a pregnancy-related disorder associated with increased cardiovascular risk for the offspring. Endothelial colony-forming cells (ECFCs) are a subset of circulating endothelial progenitor cells that participate in the formation of vasculature during development. However, the effect of preeclampsia on fetal levels of ECFCs is largely unknown. In this study, we sought to determine whether cord blood ECFC abundance and function are altered in preeclampsia. We conducted a prospective cohort study that included women with normal (n=35) and preeclamptic (n=15) pregnancies. We measured ECFC levels in the umbilical cord blood of neonates and characterized ECFC phenotype, cloning-forming ability, proliferation, and migration toward vascular endothelial growth factor-A and fibroblast growth factor-2, in vitro formation of capillary-like structures, and in vivo vasculogenic ability in immunodeficient mice. We found that the level of cord blood ECFCs was statistically lower in preeclampsia than in control pregnancies (P=0.04), a reduction that was independent of other obstetric factors. In addition, cord blood ECFCs from preeclamptic pregnancies required more time to emerge in culture than control ECFCs. However, once derived in culture, ECFC function was deemed normal and highly similar between preeclampsia and control, including the ability to form vascular networks in vivo. This study demonstrates that preeclampsia affects ECFC abundance in neonates. A reduced level of ECFCs during preeclamptic pregnancies may contribute to an increased risk of developing future cardiovascular events. © 2014 American Heart Association, Inc.

Plasmodium falciparum-infected erythrocytes induce Tissue Factor expression in endothelial cells and support the assembly of multimolecular coagulation complexes

PubMed Central

Francischetti, Ivo MB; Seydel, Karl B; Monteiro, Robson Q; Whitten, Richard O; Erexson, Cindy R; Noronha, Almério LL; Ostera, Graciela R.; Kamiza, Steve B; Molyneux, Malcolm E; Ward, Jerrold M; Taylor, Terrie E

2010-01-01

Summary Background Plasmodium falciparum malaria infects 300–500 million people every year causing 1–2 million deaths annually. Evidence of a coagulation disorder, activation of endothelial cells (EC) and increase in inflammatory cytokines are often present in malaria. Objectives We have asked whether parasitized red blood cells (pRBC) interaction with EC induces Tissue Factor expression in vitro and in vivo. The potential of phosphatidylserine-containing pRBC to support the assembly of blood coagulation complexes was also investigated. Results We demonstrate that mature forms of pRBC induce functional expression of tissue factor (TF) by endothelial cells (EC) in vitro with productive assembly of the extrinsic Xnase complex and initiation of the coagulation cascade. Late stage pRBC also support the prothrombinase and intrinsic Xnase complex formation in vitro, and may function as activated platelets in the amplification phase of the blood coagulation. Notably, postmortem brain sections obtained from P. falciparum-infected children who died from Cerebral Malaria and other causes display a consistent staining for TF in the EC. Conclusions These findings place TF expression by endothelium and the amplification of the coagulation cascade by pRBC and/or activated platelets as potentially critical steps in the pathogenesis of malaria. Furthermore, it may allow investigators to test other therapeutic alternatives targeting TF or modulators of EC function in the treatment of malaria and/or its complications. PMID:17002660

[The biological reaction of inflammation, methylglyoxal of blood plasma, functional and structural alterations in elastic type arteries at the early stage of hypertension disease].

PubMed

Titov, V N; Dmitriev, V A; Oshchepkov, E V; Balakhonova, T V; Tripoten', M I; Shiriaeva, Iu K

2012-08-01

The article deals with studying of the relationship between biologic reaction of inflammation with glycosylation reaction and content of methylglyoxal in blood serum. The positive correlation between pulse wave velocity and content of methylglyoxal, C-reactive protein in intercellular medium and malleolar brachial index value was established. This data matches the experimental results concerning involvement of biological reaction of inflammation into structural changes of elastic type arteries under hypertension disease, formation of arteries' rigidity and increase of pulse wave velocity. The arterial blood pressure is a biological reaction of hydrodynamic pressure which is used in vivo by several biological functions: biological function of homeostasis, function of endoecology, biological function of adaptation and function of locomotion. The biological reaction of hydrodynamic (hydraulic) pressure is a mode of compensation of derangement of several biological functions which results in the very high rate of hypertension disease in population. As a matter of fact, hypertension disease is a syndrome of lingering pathological compensation by higher arterial blood pressure of the biological functions derangements occurring in the distal section at the level of paracrine cenoses of cells. The arterial blood pressure is a kind of in vivo integral indicator of deranged metabolism. The essential hypertension disease pathogenically is a result of the derangement of three biological functions: biological function of homeostasis, biological function of trophology - nutrition (biological reaction of external feeding - exotrophia) and biological function of endoecology. In case of "littering" of intercellular medium in vivo with nonspecific endogenic flogogens a phylogenetically earlier activation of biological reactions of excretion, inflammation and hydrodynamic arterial blood pressure occur. In case of derangement of biological function of homeostasis, decreasing of perfusion even in single paracrine cenoses and derangement of biological function of endoecology ("purity" of intercellular medium) the only response always will be the increase of arterial blood pressure.

Post-test probability for neonatal hyperbilirubinemia based on umbilical cord blood bilirubin, direct antiglobulin test, and ABO compatibility results.

PubMed

Peeters, Bart; Geerts, Inge; Van Mullem, Mia; Micalessi, Isabel; Saegeman, Veroniek; Moerman, Jan

2016-05-01

Many hospitals opt for early postnatal discharge of newborns with a potential risk of readmission for neonatal hyperbilirubinemia. Assays/algorithms with the possibility to improve prediction of significant neonatal hyperbilirubinemia are needed to optimize screening protocols and safe discharge of neonates. This study investigated the predictive value of umbilical cord blood (UCB) testing for significant hyperbilirubinemia. Neonatal UCB bilirubin, UCB direct antiglobulin test (DAT), and blood group were determined, as well as the maternal blood group and the red blood cell antibody status. Moreover, in newborns with clinically apparent jaundice after visual assessment, plasma total bilirubin (TB) was measured. Clinical factors positively associated with UCB bilirubin were ABO incompatibility, positive DAT, presence of maternal red cell antibodies, alarming visual assessment and significant hyperbilirubinemia in the first 6 days of life. UCB bilirubin performed clinically well with an area under the receiver-operating characteristic curve (AUC) of 0.82 (95 % CI 0.80-0.84). The combined UCB bilirubin, DAT, and blood group analysis outperformed results of these parameters considered separately to detect significant hyperbilirubinemia and correlated exponentially with hyperbilirubinemia post-test probability. Post-test probabilities for neonatal hyperbilirubinemia can be calculated using exponential functions defined by UCB bilirubin, DAT, and ABO compatibility results. • The diagnostic value of the triad umbilical cord blood bilirubin measurement, direct antiglobulin testing and blood group analysis for neonatal hyperbilirubinemia remains unclear in literature. • Currently no guideline recommends screening for hyperbilirubinemia using umbilical cord blood. What is New: • Post-test probability for hyperbilirubinemia correlated exponentially with umbilical cord blood bilirubin in different risk groups defined by direct antiglobulin test and ABO blood group compatibility results. • Exponential functions can be used to calculate hyperbilirubinemia probability.

Single-cell RNA-seq reveals new types of human blood dendritic cells, monocytes and progenitors

PubMed Central

Villani, Alexandra-Chloé; Satija, Rahul; Reynolds, Gary; Sarkizova, Siranush; Shekhar, Karthik; Fletcher, James; Griesbeck, Morgane; Butler, Andrew; Zheng, Shiwei; Lazo, Suzan; Jardine, Laura; Dixon, David; Stephenson, Emily; Nilsson, Emil; Grundberg, Ida; McDonald, David; Filby, Andrew; Li, Weibo; De Jager, Philip L.; Rozenblatt-Rosen, Orit; Lane, Andrew A.; Haniffa, Muzlifah; Regev, Aviv; Hacohen, Nir

2017-01-01

Dendritic cells (DCs) and monocytes play a central role in pathogen sensing, phagocytosis and antigen presentation and consist of multiple specialized subtypes. However, their identities and interrelationships are not fully understood. Using unbiased single-cell RNA sequencing (RNA-seq) of ~2400 cells, we identified six human DCs and four monocyte subtypes in human blood. Our study reveals: a new DC subset that shares properties with plasmacytoid DCs (pDCs) but potently activates T cells, thus redefining pDCs; a new subdivision within the CD1C+ subset of DCs; the relationship between blastic plasmacytoid DC neoplasia cells and healthy DCs; and circulating progenitor of conventional DCs (cDCs). Our revised taxonomy will enable more accurate functional and developmental analyses as well as immune monitoring in health and disease. PMID:28428369

Unresolved clinical aspects and safety hazards of blood derived- EV/MV in stored blood components: From personal memory lanes to newer perspectives on the roles of EV/MV in various biological phenomena.

PubMed

Seghatchian, Jerard; Amiral, Jean

2016-08-01

Blood cells generate heterogeneous populations of vesicles that are delivered, as small-specialized packages of highly active cell fragments in blood circulation, having almost similar functional activities, as the mother cells. These so called extracellular vesicles are the essential part of an energy-dependent natural apoptotic process; hence their beneficial and harmful biological functions cannot be ignored. Evidence is accumulating, that cellular derived vesicles, originate from all viable cells including: megakaryocytes, platelets, red blood cells, white blood cells and endothelial cells, the highest in proportions from platelets. Shedding can also be triggered by pathological activation of inflammatory processes and activation of coagulation or complement pathways, or even by shear stress in the circulation. Structurally, so called MV/EV appear to be, sometimes inside-out and sometimes outside-in cell fragments having a bilayered phospholipid structure exposing coagulant-active phosphatidylserine, expressing various membrane receptors, and they serve as cell-to-cell shuttles for bioactive molecules such as lipids, growth factors, microRNAs, and mitochondria. Ex vivo processing of blood into its components, embodying centrifugation, processing by various apheresis procedures, leukoreduction, pathogen reduction, and finally storage in different media and different types of blood bags, also have major impacts on the generation and retention of MV content. These artificially generated small, but highly liable packages, together with the original pool of MVs collected from the donor, do exhibit differing biological activities, and are not inert elements and should be considered as a parameter of blood safety in haemovigilance programmes. Harmonization and consensus in sampling protocols, sample handling, processing, and assessment methods, in particular converting to full automation, are needed to achieve consensual interpretations. This review focuses on some of our past personal studies on the role of MV/EV focusing on characterization of platelet storage lesion and platelet therapy that shows the highest transfusion hazards [up to 25%], and loss of 25% platelet efficacy after various leukoreduction and validated platelet pathogen reduction treatments. The planned paths for the future of EV/MV involvement in immunological and viral/ non-viral transfusion hazards are also discussed. Whilst considerable advances made on the characterization of EV/MV, but disparity still exists between various surrogate markers, showing some subtle differences in the levels of MV/ EV & BRMs in platelet preparations, and the clinical outcome showing platelets derived by all current technologies are equivalents in vivo. One possible reason for such a disparity may be relatedto the fact that MVs, being the end products of apoptotic cells, have little specificity and clear rapidly from circulation [<6 h in thrombocytopoenia]. This makes their clinical usefulness rather short lived. The recent findings that pegylating smaller subsets of EV increases its circulatory life from <15 minutes to approximately about one hour is highly promising, in particular, for drug delivery on specific sides. Hence a promising clinical utility of EV/MV continues, as a journey without end, indeed. This manuscript is based mainly on the selected key readings listed below. Copyright © 2016. Published by Elsevier Ltd.

Synergistic effect of concanavalin A and Bu-WSA on DNA synthesis in human peripheral blood lymphocytes.

PubMed

Nitta, T; Okumura, S; Nakano, M

1985-02-01

Butanol-extracted water soluble adjuvant (Bu-WSA) obtained from Bacterionema matruchotii was not mitogenic for human peripheral blood mononuclear cells (PBM) but was capable of enhancing (3H) thymidine uptake of T cells stimulated by concanavalin A (Con A) in the presence of B cells or macrophages (M phi) in vitro. The mechanisms of the synergy of Con A and Bu-WSA were studied by using separated cell populations from PBM. Both subfractioned OKT4+ and OKT8+ cells were responsive to co-stimulation by Con A and Bu-WSA in the presence of an accessory cell population. Allogeneic B cells and M phi as well as autologous cells had helper function as accessory cells. Heavy irradiation with gamma-rays did not affect the function of the accessory cells, but previous treatment of B cells with anti-Ig serum plus complement (C) or treatment of M phi with anti-M phi serum plus C deprived them of their function. The treatment of accessory cells with anti-HLA-DR serum, regardless of the presence or absence of C, resulted in loss of their helper function. Cultures in Marbrook-type vessels showed that a mixed cell population of T cells and accessory cells in the lower chamber produced some active factor(s) after co-stimulation with Con A and Bu-WSA, and by passing through the membrane filter separating the chambers, the factor(s) enhanced the proliferation of the Con A-activated T cell population in the upper chamber. The factor(s) was presumed to be interleukin 2 (IL 2), because it supported the growth of IL 2-dependent CTLL cells. These results indicate that the synergy of Con A and Bu-WSA on the proliferative response of human PBM is due to the elevation of growth factor production from T cells stimulated by those mitogens.

Development of Cell Phone Application for Blood Glucose Self-Monitoring Based on ISO/IEEE 11073 and HL7 CCD.

PubMed

Park, Hyun Sang; Cho, Hune; Kim, Hwa Sun

2015-04-01

The objectives of this research were to develop and evaluate a cell phone application based on the standard protocol for personal health devices and the standard information model for personal health records to support effective blood glucose management and standardized service for patients with diabetes. An application was developed for Android 4.0.3. In addition, an IEEE 11073 Manager, Medical Device Encoding Rule, and Bluetooth Health Device Profile Connector were developed for standardized health communication with a glucometer, and a Continuity of Care Document (CCD) Composer and CCD Parser were developed for CCD document exchange. The developed application was evaluated by five healthcare professionals and 87 users through a questionnaire comprising the following variables: usage intention, effort expectancy, social influence, facilitating condition, perceived risk, and voluntariness. As a result of the evaluation of usability, it was confirmed that the developed application is useful for blood glucose self-monitoring by diabetic patients. In particular, the healthcare professionals stated their own views that the application is useful to observe the trends in blood glucose change through the automatic function which records a blood glucose level measured using Bluetooth function, and the function which checks accumulated records of blood glucose levels. Also, a result of the evaluation of usage intention was 3.52 ± 0.42 out of 5 points. The application developed by our research team was confirmed by the verification of healthcare professionals that accurate feedback can be provided to healthcare professionals during the management of diabetic patients or education for glucose management.

Development of Cell Phone Application for Blood Glucose Self-Monitoring Based on ISO/IEEE 11073 and HL7 CCD

PubMed Central

Park, Hyun Sang; Cho, Hune

2015-01-01

Objectives The objectives of this research were to develop and evaluate a cell phone application based on the standard protocol for personal health devices and the standard information model for personal health records to support effective blood glucose management and standardized service for patients with diabetes. Methods An application was developed for Android 4.0.3. In addition, an IEEE 11073 Manager, Medical Device Encoding Rule, and Bluetooth Health Device Profile Connector were developed for standardized health communication with a glucometer, and a Continuity of Care Document (CCD) Composer and CCD Parser were developed for CCD document exchange. The developed application was evaluated by five healthcare professionals and 87 users through a questionnaire comprising the following variables: usage intention, effort expectancy, social influence, facilitating condition, perceived risk, and voluntariness. Results As a result of the evaluation of usability, it was confirmed that the developed application is useful for blood glucose self-monitoring by diabetic patients. In particular, the healthcare professionals stated their own views that the application is useful to observe the trends in blood glucose change through the automatic function which records a blood glucose level measured using Bluetooth function, and the function which checks accumulated records of blood glucose levels. Also, a result of the evaluation of usage intention was 3.52 ± 0.42 out of 5 points. Conclusions The application developed by our research team was confirmed by the verification of healthcare professionals that accurate feedback can be provided to healthcare professionals during the management of diabetic patients or education for glucose management. PMID:25995960

Antibodies Against Immune Checkpoint Molecules Restore Functions of Tumor-Infiltrating T Cells in Hepatocellular Carcinomas.

PubMed

Zhou, Guoying; Sprengers, Dave; Boor, Patrick P C; Doukas, Michail; Schutz, Hannah; Mancham, Shanta; Pedroza-Gonzalez, Alexander; Polak, Wojciech G; de Jonge, Jeroen; Gaspersz, Marcia; Dong, Haidong; Thielemans, Kris; Pan, Qiuwei; IJzermans, Jan N M; Bruno, Marco J; Kwekkeboom, Jaap

2017-10-01

Ligand binding to inhibitory receptors on immune cells, such as programmed cell death 1 (PD-1) and cytotoxic T-lymphocyte associated protein 4 (CTLA4), down-regulates the T-cell-mediated immune response (called immune checkpoints). Antibodies that block these receptors increase antitumor immunity in patients with melanoma, non-small-cell lung cancer, and renal cell cancer. Tumor-infiltrating CD4 + and CD8 + T cells in patients with hepatocellular carcinoma (HCC) have been found to be functionally compromised. We analyzed HCC samples from patients to determine if these inhibitory pathways prevent T-cell responses in HCCs and to find ways to restore their antitumor functions. We collected HCC samples from 59 patients who underwent surgical resection from November 2013 through May 2017, along with tumor-free liver tissues (control tissues) and peripheral blood samples. We isolated tumor-infiltrating lymphocytes (TIL) and intra-hepatic lymphocytes. We used flow cytometry to quantify expression of the inhibitory receptors PD-1, hepatitis A virus cellular receptor 2 (TIM3), lymphocyte activating 3 (LAG3), and CTLA4 on CD8 + and CD4 + T cells from tumor, control tissue, and blood; we studied the effects of antibodies that block these pathways in T-cell activation assays. Expression of PD-1, TIM3, LAG3, and CTLA4 was significantly higher on CD8 + and CD4 + T cells isolated from HCC tissue than control tissue or blood. Dendritic cells, monocytes, and B cells in HCC tumors expressed ligands for these receptors. Expression of PD-1, TIM3, and LAG3 was higher on tumor-associated antigen (TAA)-specific CD8 + TIL, compared with other CD8 + TIL. Compared with TIL that did not express these inhibitory receptors, CD8 + and CD4 + TIL that did express these receptors had higher levels of markers of activation, but similar or decreased levels of granzyme B and effector cytokines. Antibodies against CD274 (PD-ligand1 [PD-L1]), TIM3, or LAG3 increased proliferation of CD8 + and CD4 + TIL and cytokine production in response to stimulation with polyclonal antigens or TAA. Importantly, combining antibody against PD-L1 with antibodies against TIM3, LAG3, or CTLA4 further increased TIL functions. The immune checkpoint inhibitory molecules PD-1, TIM3, and LAG3 are up-regulated on TAA-specific T cells isolated from human HCC tissues, compared with T cells from tumor-free liver tissues or blood. Antibodies against PD-L1, TIM3, or LAG3 restore responses of HCC-derived T cells to tumor antigens, and combinations of the antibodies have additive effects. Strategies to block PD-L1, TIM3, and LAG3 might be developed for treatment of primary liver cancer. Copyright © 2017 AGA Institute. Published by Elsevier Inc. All rights reserved.

AUTONOMIC AXONS IN THE HUMAN ENDOCRINE PANCREAS SHOW UNIQUE INNERVATION PATTERNS

PubMed Central

Rodriguez-Diaz, Rayner; Abdulreda, Midhat H.; Formoso, Alexander L.; Gans, Itai; Ricordi, Camillo; Berggren, Per-Olof; Caicedo, Alejandro

2011-01-01

SUMMARY The autonomic nervous system regulates hormone secretion from the endocrine pancreas, the islets of Langerhans, and thus impacts glucose metabolism. The parasympathetic and sympathetic nerves innervate the pancreatic islet, but the precise innervation patterns are not known, particularly in human islets. Here we demonstrate that the innervation of human islets is different from that of mouse islets and that it does not conform to existing models of autonomic control of islet function. By visualizing axons in three dimensions and quantifying axonal densities and contacts within pancreatic islets, we found that, in contrast to mouse endocrine cells, human endocrine cells are sparsely contacted by autonomic axons. Few parasympathetic cholinergic axons penetrate the human islet and the invading sympathetic fibers preferentially innervate smooth muscle cells of blood vessels located within the islet. Thus, rather than modulating endocrine cell function directly, sympathetic nerves may regulate hormone secretion in human islets by controlling local blood flow or by acting on islet regions located downstream. PMID:21723503

Direct 3D bioprinting of prevascularized tissue constructs with complex microarchitecture

PubMed Central

Zhu, Wei; Qu, Xin; Zhu, Jie; Ma, Xuanyi; Patel, Sherrina; Liu, Justin; Wang, Pengrui; Lai, Cheuk Sun Edwin; Gou, Maling; Xu, Yang; Zhang, Kang; Chen, Shaochen

2017-01-01

Living tissues rely heavily on vascular networks to transport nutrients, oxygen and metabolic waste. However, there still remains a need for a simple and efficient approach to engineer vascularized tissues. Here, we created prevascularized tissues with complex three-dimensional (3D) microarchitectures using a rapid bioprinting method – microscale continuous optical bioprinting (μCOB). Multiple cell types mimicking the native vascular cell composition were encapsulated directly into hydrogels with precisely controlled distribution without the need of sacrificial materials or perfusion. With regionally controlled biomaterial properties the endothelial cells formed lumen-like structures spontaneously in vitro. In vivo implantation demonstrated the survival and progressive formation of the endothelial network in the prevascularized tissue. Anastomosis between the bioprinted endothelial network and host circulation was observed with functional blood vessels featuring red blood cells. With the superior bioprinting speed, flexibility and scalability, this new prevascularization approach can be broadly applicable to the engineering and translation of various functional tissues. PMID:28192772

Differentiation and characterization of human pluripotent stem cell-derived brain microvascular endothelial cells.

PubMed

Stebbins, Matthew J; Wilson, Hannah K; Canfield, Scott G; Qian, Tongcheng; Palecek, Sean P; Shusta, Eric V

2016-05-15

The blood-brain barrier (BBB) is a critical component of the central nervous system (CNS) that regulates the flux of material between the blood and the brain. Because of its barrier properties, the BBB creates a bottleneck to CNS drug delivery. Human in vitro BBB models offer a potential tool to screen pharmaceutical libraries for CNS penetration as well as for BBB modulators in development and disease, yet primary and immortalized models respectively lack scalability and robust phenotypes. Recently, in vitro BBB models derived from human pluripotent stem cells (hPSCs) have helped overcome these challenges by providing a scalable and renewable source of human brain microvascular endothelial cells (BMECs). We have demonstrated that hPSC-derived BMECs exhibit robust structural and functional characteristics reminiscent of the in vivo BBB. Here, we provide a detailed description of the methods required to differentiate and functionally characterize hPSC-derived BMECs to facilitate their widespread use in downstream applications. Copyright © 2015 Elsevier Inc. All rights reserved.

Direct 3D bioprinting of prevascularized tissue constructs with complex microarchitecture.

PubMed

Zhu, Wei; Qu, Xin; Zhu, Jie; Ma, Xuanyi; Patel, Sherrina; Liu, Justin; Wang, Pengrui; Lai, Cheuk Sun Edwin; Gou, Maling; Xu, Yang; Zhang, Kang; Chen, Shaochen

2017-04-01

Living tissues rely heavily on vascular networks to transport nutrients, oxygen and metabolic waste. However, there still remains a need for a simple and efficient approach to engineer vascularized tissues. Here, we created prevascularized tissues with complex three-dimensional (3D) microarchitectures using a rapid bioprinting method - microscale continuous optical bioprinting (μCOB). Multiple cell types mimicking the native vascular cell composition were encapsulated directly into hydrogels with precisely controlled distribution without the need of sacrificial materials or perfusion. With regionally controlled biomaterial properties the endothelial cells formed lumen-like structures spontaneously in vitro. In vivo implantation demonstrated the survival and progressive formation of the endothelial network in the prevascularized tissue. Anastomosis between the bioprinted endothelial network and host circulation was observed with functional blood vessels featuring red blood cells. With the superior bioprinting speed, flexibility and scalability, this new prevascularization approach can be broadly applicable to the engineering and translation of various functional tissues. Copyright © 2017 Elsevier Ltd. All rights reserved.

Hormones and the blood-brain barrier.

PubMed

Hampl, Richard; Bičíková, Marie; Sosvorová, Lucie

2015-03-01

Hormones exert many actions in the brain, and brain cells are also hormonally active. To reach their targets in brain structures, hormones must overcome the blood-brain barrier (BBB). The BBB is a unique device selecting desired/undesired molecules to reach or leave the brain, and it is composed of endothelial cells forming the brain vasculature. These cells differ from other endothelial cells in their almost impermeable tight junctions and in possessing several membrane structures such as receptors, transporters, and metabolically active molecules, ensuring their selection function. The main ways how compounds pass through the BBB are briefly outlined in this review. The main part concerns the transport of major classes of hormones: steroids, including neurosteroids, thyroid hormones, insulin, and other peptide hormones regulating energy homeostasis, growth hormone, and also various cytokines. Peptide transporters mediating the saturable transport of individual classes of hormones are reviewed. The last paragraph provides examples of how hormones affect the permeability and function of the BBB either at the level of tight junctions or by various transporters.

Modulation of mitochondrial metabolism as a biochemical trait in blood feeding organisms: the redox vampire hypothesis redux.

PubMed

Ferreira, Caroline M; Oliveira, Matheus P; Paes, Marcia C; Oliveira, Marcus F

2018-06-01

Hematophagous organisms undergo remarkable metabolic changes during the blood digestion process, increasing fermentative glucose metabolism, and reducing respiratory rates, both consequence of functional mitochondrial remodeling. Here, we review the pathways involved in energy metabolism and mitochondrial functionality in a comparative framework across different hematophagous species, and consider how these processes regulate redox homeostasis during blood digestion. The trend across distinct species indicate that a switch in energy metabolism might represent an important defensive mechanism to avoid the potential harmful interaction of oxidants generated from aerobic energy metabolism with products derived from blood digestion. Indeed, in insect vectors, blood feeding transiently reduces respiratory rates and oxidant production, irrespective of tissue and insect model. On the other hand, a different scenario is observed in several unrelated parasite species when exposed to blood digestion products, as respiratory rates reduce and mitochondrial oxidant production increase. The emerging picture indicates that re-wiring of energy metabolism, through reduced mitochondrial function, culminates in improved tolerance to redox insults and seems to represent a key step for hematophagous organisms to cope with the overwhelming and potentially toxic blood meal. © 2018 International Federation for Cell Biology.

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling.

PubMed

Puente-Marin, Sara; Nombela, Iván; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio; Ortega-Villaizan, María Del Mar

2018-04-09

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation.

In Silico Functional Networks Identified in Fish Nucleated Red Blood Cells by Means of Transcriptomic and Proteomic Profiling

PubMed Central

Puente-Marin, Sara; Ciordia, Sergio; Mena, María Carmen; Chico, Verónica; Coll, Julio

2018-01-01

Nucleated red blood cells (RBCs) of fish have, in the last decade, been implicated in several immune-related functions, such as antiviral response, phagocytosis or cytokine-mediated signaling. RNA-sequencing (RNA-seq) and label-free shotgun proteomic analyses were carried out for in silico functional pathway profiling of rainbow trout RBCs. For RNA-seq, a de novo assembly was conducted, in order to create a transcriptome database for RBCs. For proteome profiling, we developed a proteomic method that combined: (a) fractionation into cytosolic and membrane fractions, (b) hemoglobin removal of the cytosolic fraction, (c) protein digestion, and (d) a novel step with pH reversed-phase peptide fractionation and final Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MS/MS) analysis of each fraction. Combined transcriptome- and proteome- sequencing data identified, in silico, novel and striking immune functional networks for rainbow trout nucleated RBCs, which are mainly linked to innate and adaptive immunity. Functional pathways related to regulation of hematopoietic cell differentiation, antigen presentation via major histocompatibility complex class II (MHCII), leukocyte differentiation and regulation of leukocyte activation were identified. These preliminary findings further implicate nucleated RBCs in immune function, such as antigen presentation and leukocyte activation. PMID:29642539

Enhanced Cell Capture on Functionalized Graphene Oxide Nanosheets through Oxygen Clustering.

PubMed

Bardhan, Neelkanth M; Kumar, Priyank V; Li, Zeyang; Ploegh, Hidde L; Grossman, Jeffrey C; Belcher, Angela M; Chen, Guan-Yu

2017-02-28

With the global rise in incidence of cancer and infectious diseases, there is a need for the development of techniques to diagnose, treat, and monitor these conditions. The ability to efficiently capture and isolate cells and other biomolecules from peripheral whole blood for downstream analyses is a necessary requirement. Graphene oxide (GO) is an attractive template nanomaterial for such biosensing applications. Favorable properties include its two-dimensional architecture and wide range of functionalization chemistries, offering significant potential to tailor affinity toward aromatic functional groups expressed in biomolecules of interest. However, a limitation of current techniques is that as-synthesized GO nanosheets are used directly in sensing applications, and the benefits of their structural modification on the device performance have remained unexplored. Here, we report a microfluidic-free, sensitive, planar device on treated GO substrates to enable quick and efficient capture of Class-II MHC-positive cells from murine whole blood. We achieve this by using a mild thermal annealing treatment on the GO substrates, which drives a phase transformation through oxygen clustering. Using a combination of experimental observations and MD simulations, we demonstrate that this process leads to improved reactivity and density of functionalization of cell capture agents, resulting in an enhanced cell capture efficiency of 92 ± 7% at room temperature, almost double the efficiency afforded by devices made using as-synthesized GO (54 ± 3%). Our work highlights a scalable, cost-effective, general approach to improve the functionalization of GO, which creates diverse opportunities for various next-generation device applications.

Neutrophil degranulation and immunosuppression in patients with GBM: restoration of cellular immune function by targeting arginase I.

PubMed

Sippel, Trisha R; White, Jason; Nag, Kamalika; Tsvankin, Vadim; Klaassen, Marci; Kleinschmidt-DeMasters, B K; Waziri, Allen

2011-11-15

The source of glioblastoma (GBM)-associated immunosuppression remains multifactorial. We sought to clarify and therapeutically target myeloid cell-derived peripheral immunosuppression in patients with GBM. Direct ex vivo T-cell function, serum Arginase I (ArgI) levels, and circulating myeloid lineage populations were compared between patients with GBM and normal donors or patients with other intracranial tumors. Immunofunctional assays were conducted using bulk and sorted cell populations to explore the potential transfer of myeloid cell-mediated immunosuppression and to identify a potential mechanism for these effects. ArgI-mediated immunosuppression was therapeutically targeted in vitro through pharmacologic inhibition or arginine supplementation. We identified a significantly expanded population of circulating, degranulated neutrophils associated with elevated levels of serum ArgI and decreased T-cell CD3ζ expression within peripheral blood from patients with GBM. Sorted CD11b(+) cells from patients with GBM were found to markedly suppress normal donor T-cell function in coculture, and media harvested from mitogen-stimulated GBM peripheral blood mononuclear cell (PBMC) or GBM-associated mixed lymphoid reactions showed ArgI levels that were significantly higher than controls. Critically, T-cell suppression in both settings could be completely reversed through pharmacologic ArgI inhibition or with arginine supplementation. These data indicate that peripheral cellular immunosuppression in patients with GBM is associated with neutrophil degranulation and elevated levels of circulating ArgI, and that T-cell function can be restored in these individuals by targeting ArgI. These data identify a novel pathway of GBM-mediated suppression of cellular immunity and offer a potential therapeutic window for improving antitumor immunity in affected patients.

Neutrophils are immune cells preferentially targeted by retinoic acid in elderly subjects

PubMed Central

2010-01-01

Background The immune system gradually deteriorates with age and nutritional status is a major factor in immunosenescence. Of the many nutritional factors implicated in age-related immune dysfunction, vitamin A may be a good candidate, since vitamin A concentrations classically decrease during aging whereas it may possess important immunomodulatory properties via its active metabolites, the retinoic acids. This prompted us to investigate the immune response induced by retinoids in adults and elderly healthy subjects. Before and after oral supplementation with 13cis retinoic acid (0.5 mg/kg/day during 28 days), whole blood cells were phenotyped, and functions of peripheral blood mononuclear cells (PBMC) and polymorphonuclear cells (PMN) were investigated by flow cytometry and ELISA tests. Results In both young adults (n = 20, 25 ± 4 years) and older subjects (n = 20, 65 ± 4 years), retinoic acid supplementation had no effect on the distribution of leukocyte subpopulations or on the functions of PBMC (Il-2 and sIl-2R production, membrane expression of CD25). Concerning PMN, retinoic acid induced an increase in both spontaneous migration and cell surface expression of CD11b in the two different age populations, whereas bactericidal activity and phagocytosis remained unchanged. Conclusions We demonstrated that retinoic acid induces the same intensity of immune response between adult and older subjects, and more specifically affects PMN functions, i.e. adhesion and migration, than PBMC functions. PMID:20727130

Mitochondrial function and apoptosis of peripheral mononuclear cells (PBMCs) in the HIV infected patients.

PubMed

Bociąga-Jasik, Monika; Góralska, Joanna; Polus, Anna; Śliwa, Agnieszka; Gruca, Anna; Raźny, Urszula; Zdzienicka, Anna; Garlicki, Aleksander; Mach, Tomasz; Dembińska-Kieć, Aldona

2013-06-01

HIV infection results in the development of immunodeficiency mainly due to the apoptosis of infected and by stander CD4 cells. The aim of the study was to follow the mitochondrial dependent pathway of apoptosis, one of the suggested mechanisms of above process. The inner mitochondrial membrane potential (MMP), Adenosine-5'-triphosphate (ATP) generation, apoptosis and necrosis markers of peripheral mononuclear cells (PBMCs) were compared in HIV infected patients and HIV negative control group. The correlation of blood viral load, TNFα concentration, CD4 cells count and duration of ARV therapy was considered. Additionally, group of HIV infected ARV-naive patients was involved for the follow-up study and the effects of one year of ARV therapy on measured parameters were studied. PBMCs of HIV infected individuals (especially without ARV therapy) demonstrated lower MMP and ATP generation and higher percentage of apoptotic/necrotic PBMCs. Correlation between blood TNFα level and mitochondrial dysfunction was observed. The first months of ARV therapy resulted in most significant restoration of mitochondrial function and living PBMCs count. HIV infection and ARV therapy have significant impact on mitochondrial function and apoptosis of PBMCs. They are driven by abnormal mitochondrial function apoptosis of immune cells which seems to be the key element leading to immunosuppression, thus an early intervention in this process by therapy can be beneficial for symptomatology of HIV infected patients.

Haematopoiesis: “living in the shadow” of stem cell differentiation.

PubMed

Jankowski, M; Dyszkiewicz-Konwińska, M; Magas, M; Skorupski, M; Gorecki, G; Bukowska, D; Antosik, P; Jeseta, M; Bruska, M; Nowicki, M; Zabel, M; Kempisty, B

2018-01-01

Haematopoiesis is one of the most well understood stem-cell associated processes. It is a process in which pluripotent hematopoietic stem cells (HSCs) self-proliferate and differentiate into all types of blood cells. The process takes place in marrow of the flat bones in adults, however its location changes several times through embryonic and foetal development. Given the broad range of blood cells and the major differences in their build and function, together with the fact that their numbers need to be maintained within relatively narrow margins in order to maintain homeostasis despite changing environmental conditions, makes the whole process of haematopoiesis highly regulated and depending on a variety of growth factors. When influenced by those, HSCs undergo several irreversible steps, with every next one committing them to an even more specialised fate, ending with all the specific types of mostly short-lived blood cells, that are unable to proliferate on their own and need constant replenishment from the HSC pool. Because the process of haematopoiesis is the only source of all the members of the group of cells performing a range of highly important roles in functioning of the organism, significant damage to the underlying stem cells can cause a range of severe diseases. Many treatments are suggested for managing their symptoms or slowing progress, with bone marrow transplant being one of the only ones that offer possible permanent solution and, despite being a relatively risky procedure, is being widely performed, with the methods constantly improving in order to achieve progressively better results in both treatability and survivability of the patients.

Functional properties of cells obtained from human cord blood CD34+ stem cells and mouse cardiac myocytes in coculture.

PubMed

Orlandi, Alessia; Pagani, Francesca; Avitabile, Daniele; Bonanno, Giuseppina; Scambia, Giovanni; Vigna, Elisa; Grassi, Francesca; Eusebi, Fabrizio; Fucile, Sergio; Pesce, Maurizio; Capogrossi, Maurizio C

2008-04-01

Prior in vitro studies suggested that different types of hematopoietic stem cells may differentiate into cardiomyocytes. The present work examined whether human CD34(+) cells from the human umbilical cord blood (hUCB), cocultured with neonatal mouse cardiomyocytes, acquire the functional properties of myocardial cells and express human cardiac genes. hUCB CD34(+) cells were cocultured onto cardiomyocytes following an infection with a lentivirus-encoding enhanced green fluorescent protein (EGFP). After 7 days, mononucleated EGFP(+) cells were tested for their electrophysiological features by patch clamp and for cytosolic [Ca(2+)] ([Ca(2+)](i)) homeostasis by [Ca(2+)](i) imaging of X-rhod1-loaded cells. Human Nkx2.5 and GATA-4 expression was examined in cocultured cell populations by real-time RT-PCR. EGFP(+) cells were connected to surrounding cells by gap junctions, acquired electrophysiological properties similar to those of cardiomyocytes, and showed action potential-associated [Ca(2+)](i) transients. These cells also exhibited spontaneous sarcoplasmic reticulum [Ca(2+)](i) oscillations and the associated membrane potential depolarization. However, RT-PCR of both cell populations showed no upregulation of human-specific cardiac genes. In conclusion, under our experimental conditions, hUCB CD34(+) cells cocultured with murine cardiomyocytes formed cells that exhibited excitation-contraction coupling features similar to those of cardiomyocytes. However, the expression of human-specific cardiac genes was undetectable by RT-PCR.

[Impact of oxygen toxic action on the erythrocyte membrane and possibility of estimating central nervous system function disturbances].

PubMed

Belić, Branislava; Cincović, Marko R

2011-07-01

BACKGROUND/AIM; Prolonged exposure to hyperbaric oxygen leads to changes of erythrocytes shape as a consequence of toxic effects of oxygen on the erythrocyte membrane. The aim of this study was to examine the association between occurance of pathological forms of erythrocytes at different time from the start of hyperbaric oxygenation and the moment of convulsions occurrence, an interrelationship of different pathological forms of erythrocytes during exposure to hyperbaric oxygenation, as well as the correlation between the presence of ruptured erythrocytes and function of central nervous system (CNS) after completion of hyperbaric treatment. Sixty laboratory mice, Mus musculus, were exposed to the wholly-oxygen pressure of 3.5 absolute atmospheres (ATA). Blood was collected at the 32nd, 34th, 36th, 38th and 40th minutes after the exposure to oxygen. Pathological forms of erythrocytes were examined by electron microscopy. A moment of convulsions occurrence was registered in all animals. After decompression neurological examinations of experimental animals were perfomed. The Pearson's coefficient of correlation, and linear regression equations for the parameters outlined in the aim of the study were calculated. Hyperbaric oxygen caused damages of erythrocytes at the 34th minute after beginning of the treatment. Various forms of abnormal red blood cells occured, and immediately before the occurrence of irreversible changes (erythrocyte membrane rupture) echinocyte shape was dominated. A significant correlation between the number of damaged red blood cells at 34th minute and their number at the 36th, 38th and 40th minute was found. Convulsions were diagnosed significantly earlier in mice with a greater number of damaged red blood cells (p < 0.01). There was a negative correlation between the number of irreversiblly damaged red blood cells (ruptured) at the 40th minute and neurological score in the studied animals (p < 0.05). The analysis of altered erythrocytes during hyperbaric oxygenation could predict a moment of seizures occurrence, and therefore the duration of the therapy with hyperbaric oxygen. Ehinocytes indicate impending rupture of red blood cells and a possible occurrence of seizures. An increased number of ruptured red blood cells may also even indicate the potential burden of CNS after cessation of hyperbaric oxygenation.