Lipopolysaccharide-sensitive H+ current in dendritic cells

2012 ◽  
Vol 303 (2) ◽  
pp. C204-C212 ◽  
Author(s):  
Kalina Szteyn ◽  
Wenting Yang ◽  
Evi Schmid ◽  
Florian Lang ◽  
Ekaterina Shumilina
Keyword(s):  
Dendritic Cells ◽  
Reversal Potential ◽  
Charge Compensation ◽  
Outer Wall ◽  
Functional Expression ◽  
Lymphoid Tissues ◽  
Mouse Bone Marrow ◽  
Ros Production ◽  
Whole Cell Patch Clamp ◽  
Antigen Presenting

Dendritic cells (DCs) are the most potent antigen-presenting cells equipped to transport antigens from the periphery to lymphoid tissues and to present them to T cells. Ligation of Toll-like receptor 4 (TLR4), expressed on the DC surface, by lipopolysaccharides (LPS), elements of the Gram-negative bacteria outer wall, induces DC maturation. Initial steps of maturation include stimulation of antigen endocytosis and enhanced reactive oxygen species (ROS) production with eventual downregulation of endocytic capacity in fully matured DCs. ROS production depends on NADPH oxidase (NOX2), the activity of which requires continuous pH and charge compensation. The present study demonstrates, for the first time, the functional expression of voltage-gated proton (Hv1) channels in mouse bone marrow-derived DCs. In whole cell patch-clamp experiments, we recorded Zn2+ (50 μM)-sensitive outwardly rectifying currents activated upon depolarization, which were highly selective for H+, with the reversal potential shift of 38 mV per pH unit. The threshold voltage of activation ( Vthreshold) was dependent on the pH gradient and was close to the empirically predicted Vthreshold for the Hv1 currents. LPS (1 μg/ml) had bimodal effects on Hv1 channels: acute LPS treatment increased Hv1 channel activity, whereas 24 h of LPS incubation significantly inhibited Hv1 currents and decreased ROS production. Activation of H+ currents by acute application of LPS was abolished by PKC inhibitor GFX (10 nM). According to electron current measurements, acute LPS application was associated with increased NOX2 activity.

Effect of dexamethasone on Na+/Ca2+exchanger in dendritic cells

2011 ◽  
Vol 300 (6) ◽  
pp. C1306-C1313 ◽  
Author(s):  
Nicole Heise ◽  
Ekaterina Shumilina ◽  
Meerim K. Nurbaeva ◽  
Evi Schmid ◽  
Kalina Szteyn ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Outward Current ◽  
Transcript Level ◽  
Inhibitory Effect ◽  
Mouse Bone Marrow ◽  
Glucocorticoid Treatment ◽  
Whole Cell Patch Clamp ◽  
Plasmic Reticulum ◽  
Tnf Α ◽  
Antigen Presenting

Ca+-dependent signaling regulates the function of dendritic cells (DCs), antigen-presenting cells linking innate and adaptive immunity. The activity of DCs is suppressed by glucocorticoids, potent immunosuppressive hormones. The present study explored whether the glucocorticoid dexamethasone influences the cytosolic Ca2+concentration ([Ca2+]i) in DCs. To this end, DCs were isolated from mouse bone marrow. According to fura-2 fluorescence, exposure of DCs to lipopolysaccharide (LPS, 100 ng/ml) increased [Ca2+]i, an effect significantly blunted by overnight incubation with 10 nM dexamethasone before LPS treatment. Dexamethasone did not affect the Ca2+content of intracellular stores, sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2 and SERCA3 expression, ryanodine receptor (RyR)1 expression, or Ca2+entry through store-operated Ca2+channels. In contrast, dexamethasone increased the transcript level and the membrane protein abundance of the Na+/Ca2+exchanger NCX3. The activity of Na+/Ca2+exchangers was assessed by removal of extracellular Na+in the presence of external Ca2+, a maneuver triggering the Ca2+influx mode. Indeed, Na+removal resulted in a rapid transient increase of [Ca2+]iand induced an outwardly directed current as measured in whole cell patch-clamp experiments. Dexamethasone significantly augmented the increase of [Ca2+]iand the outward current following removal of extracellular Na+. The NCX blocker KB-R7943 reversed the inhibitory effect of dexamethasone on LPS-induced increase in [Ca2+]i. Dexamethasone blunted LPS-induced stimulation of CD86 expression and TNF-α production, an effect significantly less pronounced in the presence of NCX blocker KB-R7943. In conclusion, our results show that glucocorticoid treatment blunts LPS-induced increase in [Ca2+]iin DCs by increasing expression and activity of Na+/Ca2+exchanger NCX3. The effect contributes to the inhibitory effect of the glucocorticoid on DC maturation.

Development of thymic and splenic dendritic cell populations from different hemopoietic precursors

Blood ◽  
2001 ◽  
Vol 98 (12) ◽  
pp. 3376-3382 ◽  
Author(s):  
Li Wu ◽  
Angela D'Amico ◽  
Hubertus Hochrein ◽  
Meredith O'Keeffe ◽  
Ken Shortman ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Dendritic Cells ◽  
Dendritic Cell ◽  
Cell Populations ◽  
Lymphoid Tissues ◽  
Hemopoietic Precursors ◽  
Surface Molecules ◽  
Splenic Dendritic Cell ◽  
Antigen Presenting ◽  
Macrophage Precursors

Abstract The antigen-presenting dendritic cells (DCs) found in mouse lymphoid tissues are heterogeneous. Several types of DCs have been identified on the basis of the expression of different surface molecules, including CD4, CD8α, and DEC-205. Previous studies by the authors showed that the mouse intrathymic lymphoid-restricted precursors (lin−c-kit+Thy-1lowCD4low) can produce DCs in the thymus and spleen upon intravenous transfer, suggesting a lymphoid origin of these DCs. In the current study, the potential for DC production by the newly identified bone marrow (BM) common lymphoid precursors (CLPs), common myeloid precursors (CMPs), and committed granulocyte and macrophage precursors was examined. It was found that both the lymphoid and the myeloid precursors had the potential to produce DCs. All the different DC populations identified in mouse thymus and spleen could be produced by all these precursor populations. However, CLPs produced predominantly the CD4−CD8α+ DCs, whereas CMPs produced similar numbers of CD4−CD8α+ and CD4+CD8α− DCs, although at different peak times. On a per cell basis, the CLPs were more potent than the CMPs at DC production, but this may have been compensated for by an excess of CMPs over CLPs in BM. Overall, this study shows that the expression of CD8α does not delineate the hemopoietic precursor origin of DCs, and the nature of the early precursors may bias but does not dictate the phenotype of the DC product.

scholarly journals Impact of Janus Kinase 3 on Cellular Ca2+ Release, Store Operated Ca2+ Entry and Na+/Ca2+ Exchanger Activity in Dendritic Cells

2015 ◽  
Vol 36 (6) ◽  
pp. 2287-2298 ◽  
Author(s):  
Jing Yan ◽  
Evi Schmid ◽  
Zohreh Hosseinzadeh ◽  
Sabina Honisch ◽  
Ekaterina Shumilina ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Cell Function ◽  
Janus Kinase ◽  
Transient Increase ◽  
Electrogenic Transport ◽  
Cell Responses ◽  
Whole Cell Patch Clamp ◽  
Janus Kinase 3 ◽  
Antigen Presenting ◽  
Sustained Increase

Background/Aims: Janus kinase 3 (JAK3), a tyrosine kinase mainly expressed in hematopoietic cells, participates in the signaling stimulating cell proliferation. The kinase is expressed in dendritic cells (DCs), antigen presenting cells involved in the initiation and regulation of antigen-specific T-cell responses. Dendritic cell function is regulated by cytosolic Ca2+ activity ([Ca2+]i). Mediators increasing [Ca2+]i in DCs include ATP and the chemokine receptor CXCR4 ligand CXCL12. The present study explored, whether JAK3 participates in the regulation of [Ca2+]i in DCs. Methods: Fura-2 fluorescence was employed to determine [Ca2+]i, and whole cell patch clamp to decipher electrogenic transport in immature DCs isolated from bone marrow of JAK3-knockout (jak3-/-) or wild-type mice (jak3+/+). Results: Without treatment, [Ca2+]i was similar in jak3-/- and jak3+/+ DCs. Addition of ATP (100 µM) was followed by transient increase of [Ca2+]i reflecting Ca2+ release from intracellular stores, an effect significantly less pronounced in jak3-/- DCs than in jak3+/+ DCs. CXCL12 administration was followed by a sustained increase of [Ca2+]i reflecting receptor operated Ca2+ entry, an effect significantly less rapid in jak3-/- DCs than in jak3+/+ DCs. In addition, the Ca2+ release-activated Ca2+ channel (CRAC) current triggered by IP3-induced Ca2+ store depletion and CXCL12 was significantly higher in DCs from jak3+/+ mice than in jak3-/- mice. Inhibition of sarcoendoplasmatic reticulum Ca2+-ATPase (SERCA) by thapsigargin (1 µM) in the absence of extracellular Ca2+ was followed by a transient increase of [Ca2+]i reflecting Ca2+ release from intracellular stores, and subsequent readdition of extracellular Ca2+ in the continued presence of thapsigargin was followed by a sustained increase of [Ca2+]i reflecting store operated Ca2+ entry (SOCE). Both, Ca2+ release from intracellular stores and SOCE were again significantly lower in jak3-/- DCs than in jak3+/+ DCs. Pretreatment of jak3+/+ DCs with JAK inhibitor WHI-P154 (22 µM, 10 minutes or 24 hours) significantly blunted both thapsigargin induced Ca2+ release and subsequent SOCE. Abrupt replacement of Na+ containing (130 mM) and Ca2+ free (0 mM) extracellular bath by Na+ free (0 mM) and Ca2+ containing (2 mM) extracellular bath increased [Ca2+]i reflecting Na+/Ca2+ exchanger activity, an effect again significantly less pronounced in jak3-/- DCs than in jak3+/+ DCs. Conclusions: JAK3 deficiency is followed by down-regulation of cytosolic Ca2+ release, receptor and store operated Ca2+ entry and Na+/Ca2+ exchanger activity in DCs.

scholarly journals miR29a and miR378b Influence CpG-Stimulated Dendritic Cells and Regulate cGAS/STING Pathway

Vaccines ◽  
2019 ◽  
Vol 7 (4) ◽  
pp. 197 ◽  
Author(s):  
Abid Ullah Shah ◽  
Yanan Cao ◽  
Naila Siddique ◽  
Jian Lin ◽  
Qian Yang
Keyword(s):  
Dendritic Cells ◽  
Adenosine Monophosphate ◽  
Cyclic Guanosine Monophosphate ◽  
Guanosine Monophosphate ◽  
Mouse Bone Marrow ◽  
Cytokine Detection ◽  
Sting Pathway ◽  
Factor Α ◽  
Antigen Presenting ◽  
And Control

The Cytosine–phosphate–guanosine (CpG) motif, which is specifically recognized intracellularly by dendritic cells (DCs), plays a crucial role in regulating the innate immune response. MicroRNAs (miRNAs) can strongly influence the antigen-presenting ability of DCs. In this study, we examine the action of miRNAs on CpG-stimulated and control DCs, as well as their effect on cyclic guanosine monophosphate-adenosine monophosphate (GMP–AMP) synthase (cGAS) and the stimulator of interferon genes (STING) signal pathway. Firstly, we selected miRNAs (miR-29a and miR-378b) based on expression in CpG-stimulated mouse bone marrow-derived dendritic cells (BMDCs). Secondly, we investigated the functions of miR-29a and miR-378b on CpG-stimulated and unstimulated BMDCs. The results showed that miR-29a and miR-378b increased expression of both the immunoregulatory DC surface markers (CD86 and CD40) and the immunosuppressive molecule CD273 by DCs. Thirdly, cytokine detection revealed that both miR-29a and miR-378b enhanced interferon-β (IFN-β) expression while suppressing tumor necrosis factor-α (TNF-α) production. Finally, our results suggest that miR-378b can bind TANK-binding kinase binding protein 1 (TBKBP1) to activate the cGAS/STING signaling pathway. By contrast, miR-29a targeted interferon regulatory factor 7 (IRF7) and promoted the expression of STING. Together, our results provide insight into the molecular mechanism of miRNA induction by CpG to regulate DC function.

scholarly journals Microbial Antigen-Presenting Extracellular Vesicles Derived from Genetically Modified Tumor Cells Promote Antitumor Activity of Dendritic Cells

Pharmaceutics ◽  
2021 ◽  
Vol 13 (1) ◽  
pp. 57
Author(s):  
Tomoko Ito ◽  
Kikuya Sugiura ◽  
Aya Hasegawa ◽  
Wakana Ouchi ◽  
Takayuki Yoshimoto ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Antitumor Activity ◽  
Tumor Cells ◽  
Extracellular Vesicles ◽  
Ternary Complex ◽  
Binary Complex ◽  
Mouse Bone Marrow ◽  
Bacterial Antigen ◽  
Tumor Vaccines ◽  
Antigen Presenting

Tumor-derived extracellular vesicles (EVs), as tumor vaccines, carry tumor-associated antigens (TAAs), and were expected to transfer TAAs to antigen-presenting cells. However, treatment with tumor-derived EVs exhibited no obvious antitumor effect on the established tumors, likely due to their immuno-suppressive functions, and also to the poor immunogenicity of TAAs. In order to improve the immune stimulating properties, EVs expressing a highly immunogenic bacterial antigen, 6 kDa early secretory antigenic target (ESAT-6), from Mycobacterium tuberculosis were prepared by genetically modifying the parent tumor cells with a plasmid coding for ESAT-6. Cultured B16 tumor cells were transfected with a ternary complex system consisting of pDNA, polyethylenimine (PEI), and chondroitin sulfate. The cells that were transfected with the ternary complex secreted EVs with a higher number of ESAT-6 epitopes than those transfected by a conventional DNA/PEI binary complex, due to the low cytotoxicity, and durable high expression efficiency of the ternary complex systems. The EVs presenting the ESAT-6 epitope (ESAT-EV) were collected and explored as immune modulatory agents. Dendritic cells (DCs) were differentiated from mouse bone marrow cells and incubated with ESAT-EV. After incubating with the EVs for one day, the DCs expressed a significantly higher level of DC maturation marker, CD86. The DCs treated with ESAT-EV showed a significantly improved antitumor activity in tumor-bearing mice.

scholarly journals TARM1 contributes to development of arthritis by activating dendritic cells through recognition of collagens

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Rikio Yabe ◽  
Soo-Hyun Chung ◽  
Masanori A. Murayama ◽  
Sachiko Kubo ◽  
Kenji Shimizu ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Bone Marrow Cells ◽  
Dendritic Cell Maturation ◽  
Mouse Bone Marrow ◽  
Gm Csf ◽  
Good Target ◽  
Antigen Presenting ◽  
Dc Maturation

AbstractTARM1 is a member of the leukocyte immunoglobulin-like receptor family and stimulates macrophages and neutrophils in vitro by associating with FcRγ. However, the function of this molecule in the regulation of the immune system is unclear. Here, we show that Tarm1 expression is elevated in the joints of rheumatoid arthritis mouse models, and the development of collagen-induced arthritis (CIA) is suppressed in Tarm1–/– mice. T cell priming against type 2 collagen is suppressed in Tarm1–/– mice and antigen-presenting ability of GM-CSF-induced dendritic cells (GM-DCs) from Tarm1–/– mouse bone marrow cells is impaired. We show that type 2 collagen is a functional ligand for TARM1 on GM-DCs and promotes DC maturation. Furthermore, soluble TARM1-Fc and TARM1-Flag inhibit DC maturation and administration of TARM1-Fc blocks the progression of CIA in mice. These results indicate that TARM1 is an important stimulating factor of dendritic cell maturation and could be a good target for the treatment of autoimmune diseases.

scholarly journals Live-Cell Microscopy Reveals That Human T Cells Primarily Respond Chemokinetically Within a CCL19 Gradient That Induces Chemotaxis in Dendritic Cells

2021 ◽  
Vol 12 ◽  
Author(s):  
Evert J. Loef ◽  
Hilary M. Sheppard ◽  
Nigel P. Birch ◽  
P. Rod Dunbar
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Dynamic Control ◽  
Human Monocyte ◽  
Lymphoid Tissues ◽  
Fluorescent Marker ◽  
Biased Random Walk ◽  
Human T Cells ◽  
Antigen Presenting ◽  
System Migration

The ability to study migratory behavior of immune cells is crucial to understanding the dynamic control of the immune system. Migration induced by chemokines is often assumed to be directional (chemotaxis), yet commonly used end-point migration assays are confounded by detecting increased cell migration that lacks directionality (chemokinesis). To distinguish between chemotaxis and chemokinesis we used the classic “under-agarose assay” in combination with video-microscopy to monitor migration of CCR7+ human monocyte-derived dendritic cells and T cells in response to a concentration gradient of CCL19. Formation of the gradients was visualized with a fluorescent marker and lasted several hours. Monocyte-derived dendritic cells migrated chemotactically towards the CCL19 gradient. In contrast, T cells exhibited a biased random walk that was largely driven by increased exploratory chemokinesis towards CCL19. This dominance of chemokinesis over chemotaxis in T cells is consistent with CCR7 ligation optimizing T cell scanning of antigen-presenting cells in lymphoid tissues.

I. Mucosal dendritic cells: their specialized role in initiating T cell responses

1999 ◽  
Vol 276 (5) ◽  
pp. G1074-G1078 ◽  
Author(s):  
Akiko Iwasaki ◽  
Brian L. Kelsall
Keyword(s):  
Dendritic Cells ◽  
T Cell ◽  
T Cell Responses ◽  
Antigen Presenting Cells ◽  
Lymphoid Tissues ◽  
Tissue Specific ◽  
Functional Studies ◽  
Cell Responses ◽  
And Function ◽  
Antigen Presenting

Dendritic cells (DCs) are the most competent antigen-presenting cells known for the induction of primary T cell responses. Functional studies of tissue-resident DCs have been impaired by the rarity of these cells in any given organ. Recent development of isolation procedures allowing extraction of highly purified fresh DC populations has made it possible to study mucosal DCs in distinct mucosa-associated lymphoid tissues. Here, we discuss several recent studies by us and others that describe the tissue-specific phenotype and function of mucosal DCs and speculate on the mechanism by which the resident DCs regulate tissue-specific T cell responses.

Expression of relB transcripts during lymphoid organ development: specific expression in dendritic antigen-presenting cells

Development ◽  
1993 ◽  
Vol 118 (4) ◽  
pp. 1221-1231 ◽  
Author(s):  
D. Carrasco ◽  
R.P. Ryseck ◽  
R. Bravo
Keyword(s):  
T Cells ◽  
Dendritic Cells ◽  
Signal Transduction Pathway ◽  
Antigen Presenting Cells ◽  
Lymphoid Organ ◽  
Lymphoid Tissues ◽  
Mouse Development ◽  
Specific Expression ◽  
Dendritic Cell Differentiation ◽  
Antigen Presenting

We have studied the expression of the relB gene during mouse development using in situ hybridization and immunocytochemical analysis. The results show that the expression of the relB gene is highly restricted to a subpopulation of cells that colonize the lymphoid tissues and that appear very late during the process of hematopoietic diversification. RNA transcripts of relB are very low or undetectable in early and late embryos. Low relB expression is observed in the thymus at late stages of embryogenesis but rapidly increases after birth. In adult lymphoid tissues, relB is detected in the medullary region of the thymus, the periarterial lymphatic sheaths of the spleen, and the deep cortex of the lymph nodes, which correspond to the regions where T cells of mature phenotype and interdigitating dendritic cells are present. Using double immunofluorescent labeling of thymic cell suspensions, we have identified the interdigitating dendritic cells as the target of RelB expression. These cells are part of a system of antigen-presenting cells that function in the induction of several immune responses, such as, tolerance, sensitization of MHC-restricted T cells, rejection of organ transplants and formation of T-dependent antibodies. Our observations indicate that RelB may play a particular role in the signal transduction pathway that regulate dendritic cell differentiation and its cellular responses.

scholarly journals Polarised expression pattern of focal contact proteins in highly motile antigen presenting dendritic cells

1999 ◽  
Vol 112 (11) ◽  
pp. 1685-1696
Author(s):  
J. Madruga ◽  
N.P. Koritschoner ◽  
S.S. Diebold ◽  
S.M. Kurz ◽  
M. Zenke
Keyword(s):  
Dendritic Cells ◽  
Expression Pattern ◽  
Molecular Mechanisms ◽  
Leading Edge ◽  
Lymphoid Tissues ◽  
Vimentin Expression ◽  
Human Dendritic Cells ◽  
Extracellular Matrix Interactions ◽  
Antigen Presenting

Dendritic cells are professional antigen presenting cells that capture antigens and migrate to lymphoid tissues to elicit specific T cell responses. Here we used an in vitro differentiation system for generating highly motile dendritic cells from chicken bone marrow progenitors by employing the conditional v-Rel estrogen receptor (ER) fusion protein v-RelER. Molecular mechanisms of dendritic cell motility were investigated. Differentiation of v-relER progenitors into dendritic cells is associated with a reduction in cell-cell and cell-extracellular matrix interactions as cells acquire motility. We demonstrate that v-relER progenitors and dendritic cells express several adhesion receptors and components of adhesion complexes. Differentiation of v-relER cells was accompanied by downregulation of focal adhesion kinase (FAK), a key molecule of adhesion complexes, but ectopic FAK expression did not affect cell adhesion and motility. Interestingly, v-relER dendritic cells exhibit a polarised expression pattern of actin and vimentin, with actin being highly concentrated at the leading edge of the cells where lamellipodia are formed. FAK, paxillin and tyrosine phosphorylated proteins are found at both poles of the cell and colocalise with actin at the leading edge, while surface beta1 integrin is confined to the uropod at the rear. CD34(+)stem cell-derived human dendritic cells also exhibited an elongated bipolar morphology, mode of migration and a polarised pattern of actin-vimentin expression similar to v-relER dendritic cells.

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