Rapamycin inhibits macropinocytosis and mannose receptor–mediated endocytosis by bone marrow–derived dendritic cells

Abstract Dendritic cells (DCs) are professional antigen-presenting cells (APCs) that use 2 major pathways for antigen uptake: constitutive macropinocytosis and mannose receptor–mediated endocytosis. Efficient endocytosis is critical for DCs to fulfill their sentinel function in immunity. We investigated the influence of the immunosuppressive macrolide rapamycin on macropinocytosis of fluorescein isothiocyanate (FITC)–albumin and mannose receptor–mediated endocytosis of FITC-dextran by murine bone marrow–derived DCs by flow cytometry. The data show that (1) at a low, physiologically relevant concentration (1 ng/mL), rapamycin impairs macropinocytosis and mannose receptor–mediated endocytosis; (2) the effects are independent of DC maturation and can be demonstrated specifically in immature CD11c+ major histocompatibility complex (MHC) class IIlo DCs by 3-color flow cytometry; (3) inhibition of endocytosis is not related to apoptotic cell death; and (4) molar excess of the structurally related molecule FK506 inhibits the actions of rapamycin. The inhibitory effects of rapamycin on DC endocytosis were confirmed in vivo. To our knowledge, this is the first report that a clinically relevant immunosuppressant inhibits DC endocytosis.

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Recombinant human interleukin-11 stimulates megakaryocytopoiesis and increases peripheral platelets in normal and splenectomized mice

Blood ◽

10.1182/blood.v81.4.901.901 ◽

1993 ◽

Vol 81 (4) ◽

pp. 901-908 ◽

Cited By ~ 115

Author(s):

TY Neben ◽

J Loebelenz ◽

L Hayes ◽

K McCarthy ◽

J Stoudemire ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Color Flow ◽

Platelet Counts ◽

In Vivo Treatment ◽

Interleukin 11 ◽

Megakaryocyte Progenitors

Abstract The effects of recombinant human interleukin-11 (rhIL-11) on in vivo mouse megakaryocytopoeisis were examined. Normal C57Bl/6 mice and splenectomized C57Bl/6 mice were treated for 7 days with 150 micrograms/kg rhIL-11 administered subcutaneously. In normal mice, peripheral platelet counts were elevated compared with vehicle-treated controls after 3 days of rhIL-11 treatment and remained elevated until day 10. Splenectomized mice treated with rhIL-11 showed elevated peripheral platelet counts that were similar in magnitude to normal rhIL-11-treated mice. However, on day 10 the platelet counts in rhIL-11- treated, splenectomized mice were no longer elevated. Analysis of bone marrow megakaryocyte ploidy by two-color flow cytometry showed an increase, relative to controls, in the percentage of 32N megakaryocytes in both normal and splenectomized animals treated with rhIL-11. In normal mice, the number of spleen megakaryocyte colony-forming cells (MEG-CFC) were increased twofold to threefold relative to controls after 3 and 7 days of rhIL-11 treatment, whereas the number of bone marrow MEG-CFC were increased only on day 7. The number of MEG-CFC in the bone marrow of rhIL-11-treated, splenectomized mice was increased twofold compared with controls on both days 3 and 7 of the study. These data show that in vivo treatment of normal or splenectomized mice with rhIL-11 increased megakaryocyte progenitors, stimulated endoreplication of bone marrow megakaryocytes, and increased peripheral platelet counts. In addition, results in splenectomized mice showed that splenic hematopoiesis was not essential for the observed increases in peripheral platelets in response to rhIL-11 administration.

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Influence of Epinastine Hydrochloride, an -Receptor Antagonist, on the Function of Mite Allergen-Pulsed Murine Bone Marrow-Derived Dendritic Cells In Vitro and In Vivo

Mediators of Inflammation ◽

10.1155/2009/738038 ◽

2009 ◽

Vol 2009 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

Ken-Zaburo Oshima ◽

Kazuhito Asano ◽

Ken-Ichi Kanai ◽

Miyuki Suzuki ◽

Harumi Suzaki

Keyword(s):

Bone Marrow ◽

Dendritic Cells ◽

Immune Responses ◽

Clinical Status ◽

Mouse Bone Marrow ◽

Dc Functions ◽

Receptor Antagonists ◽

Murine Bone Marrow

There is established concept that dendritic cells (DCs) play essential roles in the development of allergic immune responses. However, the influence of receptor antagonists on DC functions is not well defined. The aim of the present study was to examine the effect of epinastine hydrochloride (EP), the most notable histamine receptor antagonists in Japan, onDermatophagoides farinae (Der f)-pulsed mouse bone marrow-derived DCs in vitro and in vivo. EP at more than 25 ng/mL could significantly inhibit the production of IL-6, TNF- and IL-10 fromDer f-pulsed DCs, which was increased byDer fchallenge in vitro. On the other hand, EP increased the ability ofDer f-pulsed DCs to produce IL-12. Intranasal instillation ofDer f-pulsed DCs resulted in nasal eosinophilia associated with a significant increase in IL-5 levels in nasal lavage fluids.Der f-pulsed and EP-treated DCs significantly inhibited nasal eosinophila and reduced IL-5. These results indicate that EP inhibits the development of Th2 immune responses through the modulation of DC functions and results in favorable modification of clinical status of allergic diseases.

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Synergistic combination of murine bone marrow-derived dendritic cells loaded ex vivo with whole tumor lysate and systemic chemotherapy mediates antitumor immune responses in vivo

Biomedicine & Pharmacotherapy ◽

10.1016/j.biopha.2017.06.046 ◽

2017 ◽

Vol 93 ◽

pp. 286-295

Author(s):

Mohamed L. Salem ◽

Mohamed Nassef ◽

Soha Gomaa ◽

Ibrahim Essa

Keyword(s):

Bone Marrow ◽

Dendritic Cells ◽

Immune Responses ◽

Systemic Chemotherapy ◽

Ex Vivo ◽

Tumor Lysate ◽

Murine Bone Marrow ◽

Synergistic Combination

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D-mannose ameliorates autoimmune phenotypes in mouse models of lupus

10.21203/rs.3.rs-29878/v2 ◽

2020 ◽

Author(s):

Haiting Wang ◽

Xiangyu Teng ◽

Georges Abboud ◽

Wei Li ◽

Shuang Ye ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Cell Proliferation ◽

T Cells ◽

Dendritic Cells ◽

T Cell Proliferation ◽

Autoantibody Production ◽

Models Of Lupus

Abstract Background: Systemic lupus erythematosus is a disorder of immune regulation characterized by overproduction of autoantibodies. D-mannose is a C-2 epimer of glucose that exhibits immunoregulatory effects in models of autoimmune diseases, such as type 1 diabetes, induced rheumatoid arthritis, and airway inflammation. This study was conducted to evaluate the efficacy of D-mannose treatment in mouse models of lupus.Methods: The effect of D-Mannose was evaluated by flow cytometry on the in vitro activation of C57BL/6 (B6) murine bone marrow derived dendritic cells and their ability to induce antigen specific CD4+ T cell proliferation and activation. The effect of D-mannose administration in vivo on the frequency of Foxp3+ regulatory T cells in B6 mice was assessed by flow cytometry. D-mannose was administered to two models of lupus: the chronic graft-versus-host disease (cGVHD) induced model and the B6.lpr spontaneous model. Autoantibody production was measured by ELISA and immune activation by flow cytometry. Results were compared by two-tailed statistics: unpaired or paired t tests, or Mann-Whitney U tests depending on whether the data was normally distributed.Results: D-mannose inhibited the maturation of bone marrow dendritic cells and their induction of antigen-specific T cell proliferation and activation in vitro. In vivo, D-mannose increased the frequency of Foxp3+ regulatory T cells in unmanipulated control mice. In the cGVHD model of induced lupus, D-mannose treatment decreased autoantibody production, with a concomitant reduction of the frequency of effector memory and follicular helper T cells as well as germinal center B cells and plasma cells. These results were partially validated in the B6.lpr model of spontaneous lupus. Conclusion: Overall, our results suggest that D-mannose ameliorates autoimmune activation in models of lupus, at least partially due to its expansion of Treg cells, the induction of immature conventional dendritic cells and the downregulation of effector T cells activation. D-Mannose showed however a weaker immunomodulatory effect in lupus than in other autoimmune diseases.

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D-mannose ameliorates autoimmune phenotypes in mouse models of lupus

10.21203/rs.3.rs-29878/v1 ◽

2020 ◽

Author(s):

Haiting Wang ◽

Xiangyu Teng ◽

Georges Abboud ◽

Wei Li ◽

Shuang Ye ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Cell Proliferation ◽

T Cells ◽

Dendritic Cells ◽

T Cell Proliferation ◽

Autoantibody Production ◽

Models Of Lupus

Abstract Background Systemic lupus erythematosus is a disorder of immune regulation characterized by overproduction of autoantibodies. D-mannose is a C-2 epimer of glucose that exhibits immunoregulatory effects in models of autoimmune diseases, such as type 1 diabetes, induced rheumatoid arthritis, and airway inflammation. This study was conducted to evaluate the efficacy of D-mannose treatment in mouse models of lupus. Methods The effect of D-Mannose was evaluated by flow cytometry on the in vitro activation of C57BL/6 (B6) murine bone marrow derived dendritic cells and their ability to induce antigen specific CD4+ T cell proliferation and activation. The effect of D-mannose administration in vivo on the frequency of Foxp3+ regulatory T cells in B6 mice was assessed by flow cytometry. D-mannose was administered to two models of lupus: the chronic graft-versus-host disease (cGVHD) induced model and the B6.lpr spontaneous model. Autoantibody production was measured by ELISA and immune activation by flow cytometry. Results were compared by two-tailed statistics: unpaired or paired t tests, or Mann-Whitney U tests depending on whether the data was normally distributed. Results D-mannose inhibited the maturation of bone marrow dendritic cells and their induction of antigen-specific T cell proliferation and activation in vitro. In vivo, D-mannose increased the frequency of Foxp3+ regulatory T cells in unmanipulated control mice. In the cGVHD model of induced lupus, D-mannose treatment decreased autoantibody production, with a concomitant reduction of the frequency of effector memory and follicular helper T cells as well as germinal center B cells and plasma cells. These results were partially validated in the B6.lpr model of spontaneous lupus. Conclusion Overall, our results suggest that D-mannose ameliorates autoimmune activation in models of lupus, at least partially due to its expansion of Treg cells, the induction of immature conventional dendritic cells and the downregulation of effector T cells activation. D-Mannose showed however a weaker immunomodulatory effect in lupus than in other autoimmune diseases.

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Stimulation of megakaryocytopoiesis in mice by human recombinant interleukin-6

Blood ◽

10.1182/blood.v77.1.42.bloodjournal77142 ◽

1991 ◽

Vol 77 (1) ◽

pp. 42-48

Author(s):

RJ Hill ◽

MK Warren ◽

P Stenberg ◽

J Levin ◽

L Corash ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Interleukin 6 ◽

Platelet Volume ◽

Electron Microscopic ◽

Cross Sectional ◽

Color Flow ◽

Ploidy Levels ◽

Time Points

The in vivo effects of purified human recombinant interleukin-6 (IL-6) on murine megakaryocytopoiesis were examined. IL-6 was administered subcutaneously to Swiss Webster mice, followed by evaluation of bone marrow megakaryocyte ploidy, size and frequency, and median platelet volume 24, 48, and 72 hours after the initiation of IL-6 administration. In addition, bone marrow megakaryocyte morphology was examined using electron microscopy at 72 hours. IL-6 (10,000 U per subcutaneous injection) was administered three times during the first 24 hours, three times during the second 24 hours, and twice during the last 24-hour period. IL-6 bioactivity (10 U/ng) was determined using the IL-6-dependent murine hybridoma cell line B9. Megakaryocyte ploidy distribution, measured by two-color flow cytometry, demonstrated a shift in the modal ploidy class from 16N to 32N and a significant increase in the relative frequency of 64N megakaryocytes 48 and 72 hours (but not 24 hours) after initiation of IL-6 administration (cumulative doses of 60,000 and 80,000 U at 48 and 72 hours, respectively). In addition, ploidy levels were increased in animals that received a cumulative IL-6 dose of only 40,000 U (evaluated after 72 hours). The size of recognizable bone marrow megakaryocytes, determined by the cross-sectional areas of plastic embedded bone marrow megakaryocytes, was increased at the 48-hour (60,000 U IL-6) and 72- hour (80,000 U IL-6) time points. Megakaryocyte frequency, measured by flow cytometry, was unaffected at all time points and doses of IL-6. Median platelet volume, measured by electrical impedance, was not consistently altered by administration of IL-6. Electron microscopic examination of bone marrow megakaryocytes showed an increase in the proportion of megakaryocytes with a wide, peripheral, organelle- deficient zone from 20% +/- 9% (SD) in control animals to 50% +/- 7% (SD) (P less than .02) in animals that received IL-6. No changes were observed in the distribution of the demarcation membranes. IL-6 is a potent stimulator of murine megakaryocytopoiesis, in vivo, and appears to act early in megakaryocyte differentiation.

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Stimulation of megakaryocytopoiesis in mice by human recombinant interleukin-6

Blood ◽

10.1182/blood.v77.1.42.42 ◽

1991 ◽

Vol 77 (1) ◽

pp. 42-48 ◽

Cited By ~ 51

Author(s):

RJ Hill ◽

MK Warren ◽

P Stenberg ◽

J Levin ◽

L Corash ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Interleukin 6 ◽

Platelet Volume ◽

Electron Microscopic ◽

Cross Sectional ◽

Color Flow ◽

Ploidy Levels ◽

Time Points

Abstract The in vivo effects of purified human recombinant interleukin-6 (IL-6) on murine megakaryocytopoiesis were examined. IL-6 was administered subcutaneously to Swiss Webster mice, followed by evaluation of bone marrow megakaryocyte ploidy, size and frequency, and median platelet volume 24, 48, and 72 hours after the initiation of IL-6 administration. In addition, bone marrow megakaryocyte morphology was examined using electron microscopy at 72 hours. IL-6 (10,000 U per subcutaneous injection) was administered three times during the first 24 hours, three times during the second 24 hours, and twice during the last 24-hour period. IL-6 bioactivity (10 U/ng) was determined using the IL-6-dependent murine hybridoma cell line B9. Megakaryocyte ploidy distribution, measured by two-color flow cytometry, demonstrated a shift in the modal ploidy class from 16N to 32N and a significant increase in the relative frequency of 64N megakaryocytes 48 and 72 hours (but not 24 hours) after initiation of IL-6 administration (cumulative doses of 60,000 and 80,000 U at 48 and 72 hours, respectively). In addition, ploidy levels were increased in animals that received a cumulative IL-6 dose of only 40,000 U (evaluated after 72 hours). The size of recognizable bone marrow megakaryocytes, determined by the cross-sectional areas of plastic embedded bone marrow megakaryocytes, was increased at the 48-hour (60,000 U IL-6) and 72- hour (80,000 U IL-6) time points. Megakaryocyte frequency, measured by flow cytometry, was unaffected at all time points and doses of IL-6. Median platelet volume, measured by electrical impedance, was not consistently altered by administration of IL-6. Electron microscopic examination of bone marrow megakaryocytes showed an increase in the proportion of megakaryocytes with a wide, peripheral, organelle- deficient zone from 20% +/- 9% (SD) in control animals to 50% +/- 7% (SD) (P less than .02) in animals that received IL-6. No changes were observed in the distribution of the demarcation membranes. IL-6 is a potent stimulator of murine megakaryocytopoiesis, in vivo, and appears to act early in megakaryocyte differentiation.

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Recombinant human interleukin-11 stimulates megakaryocytopoiesis and increases peripheral platelets in normal and splenectomized mice

Blood ◽

10.1182/blood.v81.4.901.bloodjournal814901 ◽

1993 ◽

Vol 81 (4) ◽

pp. 901-908 ◽

Cited By ~ 1

Author(s):

TY Neben ◽

J Loebelenz ◽

L Hayes ◽

K McCarthy ◽

J Stoudemire ◽

...

Keyword(s):

Bone Marrow ◽

Flow Cytometry ◽

Color Flow ◽

Platelet Counts ◽

In Vivo Treatment ◽

Interleukin 11 ◽

Megakaryocyte Progenitors

The effects of recombinant human interleukin-11 (rhIL-11) on in vivo mouse megakaryocytopoeisis were examined. Normal C57Bl/6 mice and splenectomized C57Bl/6 mice were treated for 7 days with 150 micrograms/kg rhIL-11 administered subcutaneously. In normal mice, peripheral platelet counts were elevated compared with vehicle-treated controls after 3 days of rhIL-11 treatment and remained elevated until day 10. Splenectomized mice treated with rhIL-11 showed elevated peripheral platelet counts that were similar in magnitude to normal rhIL-11-treated mice. However, on day 10 the platelet counts in rhIL-11- treated, splenectomized mice were no longer elevated. Analysis of bone marrow megakaryocyte ploidy by two-color flow cytometry showed an increase, relative to controls, in the percentage of 32N megakaryocytes in both normal and splenectomized animals treated with rhIL-11. In normal mice, the number of spleen megakaryocyte colony-forming cells (MEG-CFC) were increased twofold to threefold relative to controls after 3 and 7 days of rhIL-11 treatment, whereas the number of bone marrow MEG-CFC were increased only on day 7. The number of MEG-CFC in the bone marrow of rhIL-11-treated, splenectomized mice was increased twofold compared with controls on both days 3 and 7 of the study. These data show that in vivo treatment of normal or splenectomized mice with rhIL-11 increased megakaryocyte progenitors, stimulated endoreplication of bone marrow megakaryocytes, and increased peripheral platelet counts. In addition, results in splenectomized mice showed that splenic hematopoiesis was not essential for the observed increases in peripheral platelets in response to rhIL-11 administration.

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