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1.
Cell Rep ; 29(8): 2505-2519.e4, 2019 Nov 19.
Artigo em Inglês | MEDLINE | ID: mdl-31747616

RESUMO

Human neutrophilic granulocytes form the largest pool of innate immune cells for host defense against bacterial and fungal pathogens. The dynamic changes that accompany the metamorphosis from a proliferating myeloid progenitor cell in the bone marrow into a mature non-dividing polymorphonuclear blood cell have remained poorly defined. Using mass spectrometry-based quantitative proteomics combined with transcriptomic data, we report on the dynamic changes of five developmental stages in the bone marrow and blood. Integration of transcriptomes and proteome unveils highly dynamic and differential interactions between RNA and protein kinetics during human neutrophil development, which can be linked to functional maturation of typical end-stage blood neutrophil killing activities.

2.
Blood Adv ; 3(22): 3562-3574, 2019 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-31738831

RESUMO

Myeloid-derived suppressor cells (MDSCs) have the capacity to suppress T-cell-mediated immune responses and impact the clinical outcome of cancer, infections, and transplantation settings. Although MDSCs were initially described as bone marrow-derived immature myeloid cells (either monocytic or granulocytic MDSCs), mature neutrophils have been shown to exert MDSC activity toward T cells in ways that remain unclear. In this study, we demonstrated that human neutrophils from both healthy donors and cancer patients do not exert MDSC activity unless they are activated. By using neutrophils with genetically well-defined defects, we found that reactive oxygen species (ROS) and granule-derived constituents are required for MDSC activity after direct CD11b-dependent interactions between neutrophils and T cells. In addition to these cellular interactions, neutrophils are engaged in the uptake of pieces of T-cell membrane, a process called trogocytosis. Together, these interactions led to changes in T-cell morphology, mitochondrial dysfunction, and adenosine triphosphate depletion, as indicated by electron microscopy, mass spectrometry, and metabolic parameters. Our studies characterize the different steps by which activated mature neutrophils induce functional T-cell nonresponsiveness and irreparable cell damage.

3.
Front Immunol ; 10: 2144, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31572368

RESUMO

Whereas, neutrophils have long been considered to mainly function as efficient innate immunity killers of micro-organisms at infected sites, they are now recognized to also be involved in modulation of adaptive immune responses. Immature and mature neutrophils were reported to have the capacity to suppress T cell-mediated immune responses as so-called granulocyte-myeloid-derived suppressor cells (g-MDSCs), and thereby affect the clinical outcome of cancer patients and impact the chronicity of microbial infections or rejection reactions in organ transplantation settings. These MDSCs were at first considered to be immature myeloid cells that left the bone marrow due to disease-specific signals. Current studies show that also mature neutrophils can exert suppressive activity. In this study we investigated in a robust T cell suppression assay whether immature CD11b+ myeloid cells were capable of MDSC activity comparable to mature fully differentiated neutrophils. We compared circulating neutrophils with myeloid cell fractions from the bone marrow at different differentiation stages. Our results indicate that functional MDSC activity is only becoming detectable at the final stage of differentiation, depending on the procedure of cell isolation. The MDSC activity obtained during neutrophil maturation correlated with the induction of the well-known highly mobile and toxic effector functions of the circulating neutrophil. Although immature neutrophils have been suggested to be increased in the circulation of cancer patients, we show here that immature neutrophils are not efficient in suppressing T cells. This suggests that the presence of immature neutrophils in the bloodstream of cancer patients represent a mere association or may function as a source of mature neutrophils in the tumor environment but not a direct cause of enhanced MDSC activity in cancer.

5.
Cell Rep ; 24(10): 2784-2794, 2018 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-30184510

RESUMO

Neutrophils are short-lived blood cells that play a critical role in host defense against infections. To better comprehend neutrophil functions and their regulation, we provide a complete epigenetic overview, assessing important functional features of their differentiation stages from bone marrow-residing progenitors to mature circulating cells. Integration of chromatin modifications, methylation, and transcriptome dynamics reveals an enforced regulation of differentiation, for cellular functions such as release of proteases, respiratory burst, cell cycle regulation, and apoptosis. We observe an early establishment of the cytotoxic capability, while the signaling components that activate these antimicrobial mechanisms are transcribed at later stages, outside the bone marrow, thus preventing toxic effects in the bone marrow niche. Altogether, these data reveal how the developmental dynamics of the chromatin landscape orchestrate the daily production of a large number of neutrophils required for innate host defense and provide a comprehensive overview of differentiating human neutrophils.

6.
J Allergy Clin Immunol ; 142(4): 1285-1296, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29477724

RESUMO

BACKGROUND: The genetic cause of primary immunodeficiency disease (PID) carries prognostic information. OBJECTIVE: We conducted a whole-genome sequencing study assessing a large proportion of the NIHR BioResource-Rare Diseases cohort. METHODS: In the predominantly European study population of principally sporadic unrelated PID cases (n = 846), a novel Bayesian method identified nuclear factor κB subunit 1 (NFKB1) as one of the genes most strongly associated with PID, and the association was explained by 16 novel heterozygous truncating, missense, and gene deletion variants. This accounted for 4% of common variable immunodeficiency (CVID) cases (n = 390) in the cohort. Amino acid substitutions predicted to be pathogenic were assessed by means of analysis of structural protein data. Immunophenotyping, immunoblotting, and ex vivo stimulation of lymphocytes determined the functional effects of these variants. Detailed clinical and pedigree information was collected for genotype-phenotype cosegregation analyses. RESULTS: Both sporadic and familial cases demonstrated evidence of the noninfective complications of CVID, including massive lymphadenopathy (24%), unexplained splenomegaly (48%), and autoimmune disease (48%), features prior studies correlated with worse clinical prognosis. Although partial penetrance of clinical symptoms was noted in certain pedigrees, all carriers have a deficiency in B-lymphocyte differentiation. Detailed assessment of B-lymphocyte numbers, phenotype, and function identifies the presence of an increased CD21low B-cell population. Combined with identification of the disease-causing variant, this distinguishes between healthy subjects, asymptomatic carriers, and clinically affected cases. CONCLUSION: We show that heterozygous loss-of-function variants in NFKB1 are the most common known monogenic cause of CVID, which results in a temporally progressive defect in the formation of immunoglobulin-producing B cells.

7.
J Med Genet ; 55(3): 166-172, 2018 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-29331982

RESUMO

BACKGROUND: Mutations in the NCF1 gene that encodes p47phox, a subunit of the NADPH oxidase complex, cause chronic granulomatous disease (CGD). In Kavkazi Jews, a c.579G>A (p.Trp193Ter) mutation in NCF1 is frequently found, leading to CGD. The same mutation is found in about 1% of Ashkenazi Jews, although Ashkenazi CGD patients with this mutation have never been described. METHODS: We used Sanger sequencing, multiplex ligation-dependent probe amplification (MLPA), gene scan analysis and Ion Torrent Next Generation Sequencing for genetic analysis, and measured NADPH oxidase activity and p47phox expression. RESULTS: In an Ashkenazi couple expecting a baby, both parents were found to be heterozygotes for this mutation, as was the fetus. However, segregation analysis in the extended family was consistent with the fetus inheriting both carrier alleles from the parents. MLPA indicated four complete NCF1 genes in the fetus and three in each parent. Gene sequencing confirmed these results. Analysis of fetal leucocytes obtained by cordocentesis revealed substantial oxidase activity with three different assays, which was confirmed after birth. In six additional Ashkenazi carriers of the NCF1 c.579G>A mutation, we found five individuals with three complete NCF1 genes of which one was mutated (like the parents), and one individual with in addition a fusion gene of NCF1 with a pseudogene. CONCLUSION: These results point to the existence of a 'false-carrier' state in Ashkenazi Jews and have wide implications regarding pre-pregnancy screening in this and other population groups.

8.
Endocr Connect ; 6(8): 731-740, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29101248

RESUMO

Innate immune cells have recently been identified as novel thyroid hormone (TH) target cells in which intracellular TH levels appear to play an important functional role. The possible involvement of TH receptor alpha (TRα), which is the predominant TR in these cells, has not been studied to date. Studies in TRα0/0 mice suggest a role for this receptor in innate immune function. The aim of this study was to determine whether TRα affects the human innate immune response. We assessed circulating interleukin-8 concentrations in a cohort of 8 patients with resistance to TH due to a mutation of TRα (RTHα) and compared these results to healthy controls. In addition, we measured neutrophil and macrophage function in one of these RTHα patients (mutation D211G). Circulating interleukin-8 levels were elevated in 7 out of 8 RTHα patients compared to controls. These patients harbor different mutations, suggesting that this is a general feature of the syndrome of RTHα. Neutrophil spontaneous apoptosis, bacterial killing, NAPDH oxidase activity and chemotaxis were unaltered in cells derived from the RTHαD211G patient. RTHα macrophage phagocytosis and cytokine induction after LPS treatment were similar to results from control cells. The D211G mutation did not result in clinically relevant impairment of neutrophil or pro-inflammatory macrophage function. As elevated circulating IL-8 is also observed in hyperthyroidism, this observation could be due to the high-normal to high levels of circulating T3 found in patients with RTHα.

9.
Sci Immunol ; 2(10)2017 Apr 28.
Artigo em Inglês | MEDLINE | ID: mdl-28626833

RESUMO

Bloodstream infection is a hallmark of sepsis, a medically emergent condition requiring rapid treatment. However, upregulation of host defense proteins through toll-like receptors and NFκB requires hours after endotoxin detection. Using confocal pulmonary intravital microscopy, we identified that the lung provides a TLR4-Myd88-and abl tyrosine kinase-dependent niche for immediate CD11b-dependent neutrophil responses to endotoxin and Gram-negative bloodstream pathogens. In an in vivo model of bacteremia, neutrophils crawled to and rapidly phagocytosed Escherichia coli sequestered to the lung endothelium. Therefore, the lung capillaries provide a vascular defensive niche whereby endothelium and neutrophils cooperate for immediate detection and capture of disseminating pathogens.

11.
J Immunol ; 196(3): 1272-83, 2016 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-26718340

RESUMO

Neutrophils are known to play a pivotal role in the host defense against Aspergillus infections. This is illustrated by the prevalence of Aspergillus infections in patients with neutropenia or phagocyte functional defects, such as chronic granulomatous disease. However, the mechanisms by which human neutrophils recognize and kill Aspergillus are poorly understood. In this work, we have studied in detail which neutrophil functions, including neutrophil extracellular trap (NET) formation, are involved in the killing of Aspergillus fumigatus conidia and hyphae, using neutrophils from patients with well-defined genetic immunodeficiencies. Recognition of conidia involves integrin CD11b/CD18 (and not dectin-1), which triggers a PI3K-dependent nonoxidative intracellular mechanism of killing. When the conidia escape from early killing and germinate, the extracellular destruction of the Aspergillus hyphae needs opsonization by Abs and involves predominantly recognition via Fcγ receptors, signaling via Syk, PI3K, and protein kinase C to trigger the production of toxic reactive oxygen metabolites by the NADPH oxidase and myeloperoxidase. A. fumigatus induces NET formation; however, NETs did not contribute to A. fumigatus killing. Thus, our findings reveal distinct killing mechanisms of Aspergillus conidia and hyphae by human neutrophils, leading to a comprehensive insight in the innate antifungal response.


Assuntos
Aspergilose/imunologia , Aspergillus fumigatus/imunologia , Hifas/imunologia , Neutrófilos/imunologia , Esporos Fúngicos/imunologia , Citotoxicidade Imunológica/imunologia , Armadilhas Extracelulares/imunologia , Humanos , Imunidade Inata , Síndromes de Imunodeficiência/imunologia , Microscopia Confocal , Fagócitos/imunologia
12.
Haematologica ; 101(5): 587-96, 2016 05.
Artigo em Inglês | MEDLINE | ID: mdl-26802050

RESUMO

Granulocyte transfusions are used to treat neutropenic patients with life-threatening bacterial or fungal infections that do not respond to anti-microbial drugs. Donor neutrophils that have been mobilized with granulocyte-colony stimulating factor (G-CSF) and dexamethasone are functional in terms of antibacterial activity, but less is known about their fungal killing capacity. We investigated the neutrophil-mediated cytotoxic response against C. albicans and A. fumigatus in detail. Whereas G-CSF/dexamethasone-mobilized neutrophils appeared less mature as compared to neutrophils from untreated controls, these cells exhibited normal ROS production by the NADPH oxidase system and an unaltered granule mobilization capacity upon stimulation. G-CSF/dexamethasone-mobilized neutrophils efficiently inhibited A. fumigatus germination and killed Aspergillus and Candida hyphae, but the killing of C. albicans yeasts was distinctly impaired. Following normal Candida phagocytosis, analysis by mass spectrometry of purified phagosomes after fusion with granules demonstrated that major constituents of the antimicrobial granule components, including major basic protein (MBP), were reduced. Purified MBP showed candidacidal activity, and neutrophil-like Crisp-Cas9 NB4-KO-MBP differentiated into phagocytes were impaired in Candida killing. Together, these findings indicate that G-CSF/dexamethasone-mobilized neutrophils for transfusion purposes have a selectively impaired capacity to kill Candida yeasts, as a consequence of an altered neutrophil granular content.


Assuntos
Candida albicans/imunologia , Citotoxicidade Imunológica , Granulócitos/imunologia , Transfusão de Leucócitos , Viabilidade Microbiana/imunologia , Biomarcadores , Degranulação Celular/efeitos dos fármacos , Degranulação Celular/imunologia , Grânulos Citoplasmáticos/imunologia , Grânulos Citoplasmáticos/metabolismo , Dexametasona/farmacologia , Fator Estimulador de Colônias de Granulócitos/farmacologia , Granulócitos/efeitos dos fármacos , Granulócitos/metabolismo , Granulócitos/microbiologia , Humanos , Imunofenotipagem , NADPH Oxidases/metabolismo , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/metabolismo , Neutrófilos/microbiologia , Fagocitose/efeitos dos fármacos , Fagocitose/imunologia , Fagossomos/imunologia , Fagossomos/microbiologia
14.
Blood ; 124(4): 590-7, 2014 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-24948657

RESUMO

Invasive fungal infections, accompanied by high rates of mortality, represent an increasing problem in medicine. Neutrophils are the major effector immune cells in fungal killing. Based on studies with neutrophils from patients with defined genetic defects, we provide evidence that human neutrophils use 2 distinct and independent phagolysosomal mechanisms to kill Candida albicans. The first mechanism for the killing of unopsonized C albicans was found to be dependent on complement receptor 3 (CR3) and the signaling proteins phosphatidylinositol-3-kinase and caspase recruitment domain-containing protein 9 (CARD9), but was independent of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity. The second mechanism for the killing of opsonized C albicans was strictly dependent on Fcγ receptors, protein kinase C (PKC), and reactive oxygen species production by the NADPH oxidase system. Each of the 2 pathways of Candida killing required Syk tyrosine kinase activity, but dectin-1 was dispensable for both of them. These data provide an explanation for the variable clinical presentation of fungal infection in patients suffering from different immune defects, including dectin-1 deficiency, CARD9 deficiency, or chronic granulomatous disease.


Assuntos
Candida albicans/imunologia , Candidíase/prevenção & controle , Imunidade Inata/imunologia , Neutrófilos/imunologia , Candida albicans/crescimento & desenvolvimento , Candidíase/imunologia , Candidíase/microbiologia , Células Cultivadas , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lectinas Tipo C/antagonistas & inibidores , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , NADPH Oxidases/antagonistas & inibidores , NADPH Oxidases/genética , NADPH Oxidases/metabolismo , Fagocitose , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/genética , Proteína Quinase C/metabolismo , Proteínas Tirosina Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptores de IgG/antagonistas & inibidores , Receptores de IgG/genética , Receptores de IgG/metabolismo , Quinase Syk
15.
J Neuroinflammation ; 11: 23, 2014 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-24485070

RESUMO

BACKGROUND: In neuroinflammatory diseases, macrophages can play a dual role in the process of tissue damage, depending on their activation status (M1 / M2). M1 macrophages are considered to exert damaging effects to neurons, whereas M2 macrophages are reported to aid regeneration and repair of neurons. Their migration within the central nervous system may be of critical importance in the final outcome of neurodegeneration in neuroinflammatory diseases e.g. multiple sclerosis (MS). To provide insight into this process, we examined the migratory capacity of human monocyte-derived M1 and M2 polarised macrophages towards chemoattractants, relevant for neuroinflammatory diseases like MS. METHODS: Primary cultures of human monocyte-derived macrophages were exposed to interferon gamma and lipopolysaccharide (LPS) to evoke proinflammatory (M1) activation or IL-4 to evoke anti-inflammatory (M2) activation. In a TAXIScan assay, migration of M0, M1 and M2 towards chemoattractants was measured and quantified. Furthermore the adhesion capacity and the expression levels of integrins as well as chemokine receptors of M0, M1 and M2 were assessed. Alterations in cell morphology were analysed using fluorescent labelling of the cytoskeleton. RESULTS: Significant differences were observed between M1 and M2 macrophages in the migration towards chemoattractants. We show that M2 macrophages migrated over longer distances towards CCL2, CCL5, CXCL10, CXCL12 and C1q compared to non-activated (M0) and M1 macrophages. No differences were observed in the adhesion of M0, M1 and M2 macrophages to multiple matrix components, nor in the expression of integrins and chemokine receptors. Significant changes were observed in the cytoskeleton organization upon stimulation with CCL2, M0, M1 and M2 macrophages adopt a spherical morphology and the cytoskeleton is rapidly rearranged. M0 and M2 macrophages are able to form filopodia, whereas M1 macrophages only adapt a spherical morphology. CONCLUSIONS: Together our results indicate that the alternative activation status of macrophages promotes their migratory properties to chemoattractants relevant for neuroinflammatory diseases like MS. Conversely, classically activated, proinflammatory macrophages have reduced migratory properties. Based on our results, we postulate that the activation status of the macrophage influences the capacity of the macrophages to rearrange their cytoskeleton. This is the first step in understanding how modulation of macrophage activation affects macrophage migration in neuroinflammatory diseases like MS.


Assuntos
Movimento Celular/fisiologia , Citocinas/metabolismo , Citoesqueleto/metabolismo , Regulação da Expressão Gênica/fisiologia , Macrófagos/fisiologia , Adesão Celular , Células Cultivadas , Complemento C1q/metabolismo , Ensaio de Imunoadsorção Enzimática , Humanos , Interferon gama/farmacologia , Lipopolissacarídeos/farmacologia , Receptores de Quimiocinas/metabolismo
17.
Blood ; 122(1): 109-11, 2013 Jul 04.
Artigo em Inglês | MEDLINE | ID: mdl-23687090

RESUMO

Familial hemophagocytic lymphohistiocytosis (FHL) is caused by genetic defects in cytotoxic granule components or their fusion machinery, leading to impaired natural killer cell and/or T lymphocyte degranulation and/or cytotoxicity. This may accumulate into a life-threatening condition known as macrophage activation syndrome. STXBP2, also known as MUNC18-2, has recently been identified as the disease-causing gene in FHL type 5 (FHL-5). A role for STXBP2 in neutrophils, and for neutrophils in FHL in general, has not been documented thus far. Here, we report that FHL-5 neutrophils have a profound defect in granule mobilization, resulting in inadequate bacterial killing, in particular, of gram-negative Escherichia coli, but not of Staphylococcus aureus, which rather depends on intact reduced NAD phosphate oxidase activity. This impairment of bacterial killing may contribute to the apparent susceptibility to gastrointestinal tract inflammation in patients with FHL-5.


Assuntos
Gastroenterite/imunologia , Linfo-Histiocitose Hemofagocítica/imunologia , Proteínas Munc18/genética , Proteínas Munc18/imunologia , Neutrófilos/imunologia , Degranulação Celular/genética , Degranulação Celular/imunologia , Grânulos Citoplasmáticos/metabolismo , Grânulos Citoplasmáticos/microbiologia , Escherichia coli/imunologia , Infecções por Escherichia coli/genética , Infecções por Escherichia coli/imunologia , Feminino , Gastroenterite/genética , Predisposição Genética para Doença , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/microbiologia , Linfo-Histiocitose Hemofagocítica/genética , Linfo-Histiocitose Hemofagocítica/microbiologia , Masculino , Neutrófilos/microbiologia , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/imunologia
18.
Blood ; 121(13): 2385-92, 2013 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-23335372

RESUMO

Caspase recruitment domain-containing protein 9 (CARD9) is an adaptor molecule in the cytosol of myeloid cells, required for induction of T-helper cells producing interleukin-17 (Th17 cells) and important in antifungal immunity. In a patient suffering from Candida dubliniensis meningoencephalitis, mutations in the CARD9 gene were found to result in the loss of protein expression. Apart from the reduced numbers of CD4(+) Th17 lymphocytes, we identified a lack of monocyte-derived cytokines in response to Candida strains. Importantly, CARD9-deficient neutrophils showed a selective Candida albicans killing defect with abnormal ultrastructural phagolysosomes and outgrowth of hyphae. The neutrophil killing defect was independent of the generation of reactive oxygen species by the reduced NAD phosphate oxidase system. Taken together, this demonstrates that human CARD9 deficiency results in selective defect in the host defense against invasive fungal infection, caused by an impaired phagocyte killing.


Assuntos
Proteínas Adaptadoras de Sinalização CARD/deficiência , Proteínas Adaptadoras de Sinalização CARD/genética , Candidíase Invasiva/imunologia , Neutrófilos/imunologia , Adolescente , Encefalopatias/diagnóstico , Encefalopatias/etiologia , Encefalopatias/imunologia , Proteínas Adaptadoras de Sinalização CARD/imunologia , Candida albicans/imunologia , Candida albicans/isolamento & purificação , Candidíase Invasiva/complicações , Candidíase Invasiva/genética , Células Cultivadas , Citofagocitose/genética , Citofagocitose/imunologia , Feminino , Humanos , Imunidade Inata/genética , Imunidade Inata/fisiologia
19.
Cell Rep ; 2(4): 748-55, 2012 Oct 25.
Artigo em Inglês | MEDLINE | ID: mdl-23022485

RESUMO

The phagocyte NADPH oxidase mediates oxidative microbial killing in granulocytes and macrophages. However, because the reactive oxygen species produced by the NADPH oxidase can also be toxic to the host, it is essential to control its activity. Little is known about the endogenous mechanism(s) that limits NADPH oxidase activity. Here, we demonstrate that the myeloid-inhibitory receptor SIRPα acts as a negative regulator of the phagocyte NADPH oxidase. Phagocytes isolated from SIRPα mutant mice were shown to have an enhanced respiratory burst. Furthermore, overexpression of SIRPα in human myeloid cells prevented respiratory burst activation. The inhibitory effect required interactions between SIRPα and its natural ligand, CD47, as well as signaling through the SIRPα cytoplasmic immunoreceptor tyrosine-based inhibitory motifs. Suppression of the respiratory burst by SIRPα was caused by a selective repression of gp91(phox) expression, the catalytic component of the phagocyte NADPH oxidase complex. Thus, SIRPα can limit gp91(phox) expression during myeloid development, thereby controlling the magnitude of the respiratory burst in phagocytes.


Assuntos
Regulação da Expressão Gênica , Glicoproteínas de Membrana/metabolismo , NADPH Oxidases/metabolismo , Fagócitos/enzimologia , Receptores Imunológicos/metabolismo , Animais , Antígeno CD47/metabolismo , Diferenciação Celular , Linhagem Celular , Granulócitos/imunologia , Granulócitos/metabolismo , Humanos , Macrófagos/imunologia , Macrófagos/metabolismo , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Células Mieloides/metabolismo , NADPH Oxidase 2 , NADPH Oxidases/genética , Fagocitose , Espécies Reativas de Oxigênio/metabolismo , Receptores Imunológicos/genética , Explosão Respiratória , Transdução de Sinais
20.
Transfusion ; 51(11): 2358-66, 2011 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-21575006

RESUMO

BACKGROUND: Bioactive lipids (lysophosphatidylcholines [lysoPCs]) accumulating during storage of cell-containing blood products are thought to be causative in onset of transfusion-related acute lung injury through activation of neutrophils. LysoPCs are thought to be derived from cell membrane degradation products such as phosphatidylcholines (PC) by partial hydrolysis of PC, a process that is catalyzed by phospholipase A(2) (PLA(2) ). STUDY DESIGN AND METHODS: We investigated the underlying mechanisms of lysoPC generation and its contribution to in vitro neutrophil-priming capacity during storage of red blood cells (RBCs), platelet (PLTs) concentrates, and cell-free plasma. Blood from healthy volunteers was drawn, processed, and stored according to Sanquin Blood Bank protocols. RESULTS: Storage of RBCs in saline-adenine-glucose-mannitol (SAGM) did not result in accumulation of lysoPCs or neutrophil-priming capacity. Replacement of SAGM by plasma as RBC storage medium caused elevated lysoPC levels on Day 0, which did not further increase during storage. Cell-free plasma stored at 22°C showed accumulation of lysoPCs during storage, which was not present at 4°C. Addition of a soluble PLA(2) or cytosolic PLA(2) inhibitor did not prevent accumulation of lysoPCs in plasma. In PLTs, lysoPC accumulation during storage was plasma dependent, but lysoPCs did not explain the observed neutrophil-priming effect as preventing accumulation of lysoPCs by removing the plasma fraction did not prevent the neutrophil-priming capacity. CONCLUSION: Accumulation of lysoPCs during storage is not cell but plasma derived and storage temperature dependent and does not explain the neutrophil-priming effect of aged products.


Assuntos
Preservação de Sangue , Lisofosfatidilcolinas/metabolismo , Reação Transfusional , Lesão Pulmonar Aguda/etiologia , Adulto , Proteínas Sanguíneas/farmacologia , Humanos , Neutrófilos/fisiologia , Temperatura Ambiente
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