Increased expression of the tetraspanins CD53 and CD63 on apoptotic human neutrophils

Beinert, T.; Münzing, Silvia; Possinger, K.; Krombach, Fritz

doi:10.1002/jlb.67.3.369

Cited by 43 publications

(27 citation statements)

References 42 publications

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“…It is, therefore, likely that TIMP-1 directly acts on hematopoietic progenitors, triggering their differentiation towards the granulocytic lineage. As CD63 itself is described to regulate activation and adhesion of mature neutrophils to endothelium, 51 granule targeting and release, 52 and is up-regulated on apoptotic neutrophils, 53 TIMP-1 /CD63 interactions may have additional effects on mature neutrophils.…”

Section: Discussionmentioning

confidence: 99%

TIMP-1 signaling via CD63 triggers granulopoiesis and neutrophilia in mice

et al. 2015

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© F e r r a t a S t o r t i F o u n d a t i o nrent study, we investigated whether TIMP-1 affects neutrophil homeostasis. We show that TIMP-1 signaling via CD63 triggered granulopoiesis in the bone marrow (BM) resulting in increased systemic neutrophil blood counts. Our findings reveal a new function of TIMP-1 on immune cell homeostasis, and may provide a link to the frequently described correlation of high TIMP-1 levels with inflammatory diseases in the clinic. Methods Animal experimentsAnimal experiments were performed in compliance with the guidelines of the Tierschutzgesetz des Freistaates Bayern and approved by the Regierung von Oberbayern. For TIMP-1-secreting tumors, 2x10 6 HT1080 cells were subcutaneously injected into the nuchal fold of CD1 nu/nu mice and tumors were grown over ten days to a diameter of approximately 1 cm. Tumor size was monitored with a caliper. For adenoviral transduction, 3x10 9 ifu were intravenously injected, as previously described. 22 Recombinant TIMP-1 protein was applied in LPS-free PBS by a single intraperitoneal (i.p.) injection of 2 mg/kg, as previously described. 22 Mice were killed with CO 2 , blood was collected from the vena cava inferior using EDTA-coated syringes, and BM cells were obtained by flushing femurs and tibias with PBS. Flow cytometryAbsolute quantification of blood neutrophils occurred in BD trucount TM tubes according to the manufacturer´s instructions. Briefly, antibody-cocktail was mixed with 50 mL fresh blood in trucount TM tubes and incubated for 15 min at RT. Subsequently, 450 mL FACS TM lysing solution (BD Bioscience) were added and incubated for 30 min at RT to lyse erythrocytes and fix the sample. For staining of BM, femurs and tibias were flushed with PBS and the obtained cell suspension was stained as blood. Samples were measured on a FACS Canto TM II flow cytometer (BD) and analyzed using FlowJo software. Immunohistochemical staining of neutrophilsFor immunohistochemical analysis of BM, femurs were freed from tissue, fixed in 4% paraformaldehyde for 20 h, de-calcified in 0.5 M EDTA for five days, dehydrated and embedded into paraffin. Ly6G-positive cells were stained on 4 mm sections, as previously described. 22 Colony formation assayBone marrow cells were harvested from AdCtrl or AdT1-transduced mice and erythrocytes were removed with RBC lysis buffer (eBioscience). 1x10 4 BM cells were seeded in MethoCult TM medium (StemCell Technologies) containing 50 ng/mL murine rSCF, 10 ng/mL murine rIL-3, and 10 ng/mL murine rIL-6 into 6-well plates. Cells were incubated at 37°C for seven days and colonies per well were counted. For each mouse, analysis was performed in duplicates, and a total of 5 mice per group were analyzed. 32Dcl3 differentiation assay 32Dcl3 cells were seeded at a density of 8x10 5 cells/mL in IL3-free medium supplemented with 100 ng/mL murine G-CSF (Peptrotech) +/-1000 ng/mL rTIMP-1. Every two days, medium was replaced by fresh medium containing 50 ng/mL G-CSF +/-1000 ng/mL rTIMP-1. At the indicated time points, cells were c...

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Section: Discussionmentioning

confidence: 99%

TIMP-1 signaling via CD63 triggers granulopoiesis and neutrophilia in mice

et al. 2015

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“…Those then isolated cells were used for the anti-IgE activation of the basophilic cells. Because the CD63-positive platelets [27], monocytes [28], dendritic cells [29], apoptotic granulocytes [30], and eosinophilic granulocytes [31,32] could falsify the result, the electronic window on the FACS was cued to the basophils (red-stained cloud of dots) by adjusting the FSS/SSC settings ( fig. 2a) HLA-DR -negative cells (R2 in fig.…”

Section: Discussionmentioning

confidence: 99%

Sensitive Flow Cytometric Method to Test Basophil Activation Influenced by Homeopathic Histamine Dilutions

Lorenz¹,

Schneider

Stolz³

et al. 2003

Complement Med Res

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Objective: In an experimental setting, human basophil degranulation was triggered by anti-IgE to measure the effects from homeopathic solutions in an in-vitro cell system. A 3-color flow cytometric method with enhanced accuracy was established. As an example we looked at the influence of histamine on anti-IgE activation of basophils. Methods: Basophils were identified in the flow cytometer by their physical properties in the forward and side scatter light depiction and by gating on CD2^–, CD14^–, CD16^–, CD19^–, HLA-DR^– negative and CD123-positive cells. CD63 expression on the cell surface of the anti-IgE-activated basophils served as an activation marker. Results: With this method we were able to study basophil function of the 0.6–3.9% basophils out of the mononuclear blood cell fraction and to document their activation status upon anti-IgE activation. Optimal activation occurs at 0.6 µg/ml final anti-IgE concentration; not less than 10,000 basophils have to be counted per batch to reduce the variation of the measurement. The fixation method was able to stabilize activation for two days. After investigation and reduction of the source of measurement variability, an unequivocally inhibited basophil activation was documented in a partly optimized system with homeopathic dilutions of histamine (10^–22M, 10^–23M, 10^–24M, and 10^–25M histamine). Dilutions greater than 10^–20M histamine (Avogadro’s number 6.02 × 10²³) account for less than 1.36 molecules of histamine in the test sample, indicating a true homeopathic effect. Conclusions: This test system is adequate for studying the effects of highly diluted mediators on basophil activation by anti-IgE. The systematic application of this experimental arrangement is recommended to study the effects of homeopathic dilutions on basophils.

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“…The expression of CD53 (a tetraspanin) is elevated in both apoptotic human neutrophils 63 and in an apoptosis-sensitive B-cell mouse lymphoma cell line. 64 The lysosome-associated protein LAPTM5 was originally identified as a gene that was preferentially expressed by adult human hematopoietic tissues, 65 but a rat homologue of this gene was recently found to be up-regulated in response to neuronal apoptosis.…”

Section: Expression Profiling Of T(14;18)-positive Cell Lines 131mentioning

confidence: 99%

Microarray Analysis of B-Cell Lymphoma Cell Lines with the t(14;18)

Robetorye

Bohling

Morgan

et al. 2002

The Journal of Molecular Diagnostics

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The t(14;18) is the most common genetic alteration in follicular lymphoma, and is detectable in a subset of diffuse large B-cell lymphomas (DLBCL), resulting in over-expression of the anti-apoptotic protein BCL-2. Although the t(14;18)-induced over-expression of BCL-2 is an important step in lymphomagenesis,The t(14;18) is a frequent chromosomal alteration in nonHodgkin's lymphomas (NHL), occurring in up to 90% of follicular lymphomas 1 and 20 to 30% of diffuse large B-cell lymphomas (DLBCL). 2,3 This translocation is acquired early in B-cell development and results in juxtaposition of the bcl-2 gene on chromosome 18 with the immunoglobulin heavy-chain locus on chromosome 14, resulting deregulated expression of a structurally intact BCL-2 protein. 4 -8 BCL-2 is located in the inner mitochondrial membrane and functions as an anti-apoptotic protein that inhibits programmed cell death, 9 resulting in accumulation of B-lymphocytes by virtue of increased cell survival. 9 -11 Although the t(14;18)-induced over-expression of bcl-2 is an important step in lymphomagenesis, this alteration alone is not sufficient to produce malignant lymphoma. This has been demonstrated in in vitro gene transfer experiments using cell lines and transgenic mice studies in which forced over-expression of bcl-2 was insufficient to cause lymphoma. 12,13 Indeed, low numbers of t(14; 18)-carrying cells can be detected in benign lymphoid tissues such as follicular hyperplasias and the hyperplastic tonsils of young children. 14 These studies indicate that cumulative genetic and cellular alterations superimposed on the t(14;18) are necessary for the development of lymphoma and underscore the need for further analysis of t(14;18)-containing DLBCLs to identify additional genes involved in lymphomagenesis and progression.cDNA microarray technology allows large scale parallel analyses of gene expression and permits simultaneous comparison of the relative gene expression levels of several thousand genes in different cell types. 15,16 This approach has now been used successfully by a number

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Increased expression of the tetraspanins CD53 and CD63 on apoptotic human neutrophils

Cited by 43 publications

References 42 publications

TIMP-1 signaling via CD63 triggers granulopoiesis and neutrophilia in mice

TIMP-1 signaling via CD63 triggers granulopoiesis and neutrophilia in mice

Sensitive Flow Cytometric Method to Test Basophil Activation Influenced by Homeopathic Histamine Dilutions

Microarray Analysis of B-Cell Lymphoma Cell Lines with the t(14;18)

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