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1.
Proc Natl Acad Sci U S A ; 106(11): 4331-6, 2009 Mar 17.
Article in English | MEDLINE | ID: mdl-19246373

ABSTRACT

Efficient vaccines potentiate antibody avidity and increase T cell longevity, which confer protection against microbial lethal challenge. A vaccine strategy was established by using Lactobacillus acidophilus to deliver Bacillus anthracis protective antigen (PA) via specific dendritic cell-targeting peptides to dendritic cells (DCs), which reside in the periphery and mucosal surfaces, thus directing and regulating acquired immunity. The efficiency of oral delivery of L. acidophilus expressing a PA-DCpep fusion was evaluated in mice challenged with lethal B. anthracis Sterne. Vaccination with L. acidophilus expressing PA-DCpep induced robust protective immunity against B. anthracis Sterne compared with mice vaccinated with L. acidophilus expressing PA-control peptide or an empty vector. Additionally, serum anti-PA titers, neutralizing PA antibodies, and the levels of IgA-expressing cells were all comparable with the historical recombinant PA plus aluminum hydroxide vaccine administered s.c. Collectively, development of this strategy for oral delivery of DC-targeted antigens provides a safe and protective vaccine via a bacterial adjuvant that may potentiate mucosal immune responses against deadly pathogens.


Subject(s)
Anthrax Vaccines/immunology , Antigens, Bacterial/therapeutic use , Bacillus anthracis/immunology , Dendritic Cells/immunology , Lactobacillus acidophilus/genetics , Administration, Oral , Animals , Anthrax/prevention & control , Anthrax Vaccines/administration & dosage , Antibody Formation , Antigen Presentation , Antigens, Bacterial/administration & dosage , Antigens, Bacterial/genetics , Immunity , Mice
2.
J Virol ; 78(19): 10258-64, 2004 Oct.
Article in English | MEDLINE | ID: mdl-15367591

ABSTRACT

The appearance of virus-specific CD4(+) and/or CD8(+) T lymphocytes in peripheral blood of captive juvenile rhesus macaques (Macaca mulatta) was observed following rotavirus infection. These cell-mediated immune responses were measured following experimental or natural infection after rotavirus was isolated from stool specimens of asymptomatic animals. The virus isolated was a new strain of simian rotavirus that we named TUCH (for Tulane University and Cincinnati Children's Hospital). Restimulation of peripheral T lymphocytes by inactivated double- or triple-layered TUCH rotavirus particles containing either VP6 or VP4 and VP7 on their respective surfaces resulted in increased quantities of interleukin-6 (IL-6) and IL-12 in cell culture supernatants. Recall responses to rotavirus by CD4(+) and CD8(+) T lymphocytes were associated with accumulation of intracellular IL-6 and gamma interferon. Antigen presentation of TUCH rotavirus to lymphocytes was mediated via differentiated cultures of monocyte-derived dendritic (HLA-DR(+)) cells. This is the first report demonstrating cell-mediated immune responses to rotavirus in nonhuman primates. Further exploration of rhesus macaques in vaccine trials with human rotavirus vaccine candidates is the major objective of future studies.


Subject(s)
Rotavirus Infections/immunology , Rotavirus/immunology , T-Lymphocytes/immunology , Animals , Antigen Presentation/immunology , Antigens, Viral/immunology , CD4-Positive T-Lymphocytes/immunology , CD8-Positive T-Lymphocytes/immunology , Capsid Proteins/immunology , Cells, Cultured , Dendritic Cells/immunology , Feces/virology , Interferon-gamma/metabolism , Interleukin-12/metabolism , Interleukin-6/metabolism , Lymphocyte Activation , Macaca mulatta , Rotavirus/classification , Rotavirus/isolation & purification , T-Lymphocytes/metabolism
3.
J Exp Med ; 194(7): 1013-20, 2001 Oct 01.
Article in English | MEDLINE | ID: mdl-11581322

ABSTRACT

Langerhans cells (LCs) represent a subset of immature dendritic cells (DCs) specifically localized in the epidermis and other mucosal epithelia. As surrounding keratinocytes can produce interleukin (IL)-15, a cytokine that utilizes IL-2Rgamma chain, we analyzed whether IL-15 could skew monocyte differentiation into LCs. Monocytes cultured for 6 d with granulocyte/macrophage colony-stimulating factor (GM-CSF) and IL-15 differentiate into CD1a(+)HLA-DR(+)CD14(-)DCs (IL15-DCs). Agents such as lipopolysaccharide (LPS), tumor necrosis factor (TNF)alpha, and CD40L induce maturation of IL15-DCs to CD83(+), DC-LAMP(+) cells. IL15-DCs are potent antigen-presenting cells able to induce the primary (mixed lymphocyte reaction [MLR]) and secondary (recall responses to flu-matrix peptide) immune responses. As opposed to cultures made with GM-CSF/IL-4 (IL4-DCs), a proportion of IL15-DCs expresses LC markers: E-Cadherin, Langerin, and CC chemokine receptor (CCR)6. Accordingly, IL15-DCs, but not IL4-DCs, migrate in response to macrophage inflammatory protein (MIP)-3alpha/CCL20. However, IL15-DCs cannot be qualified as "genuine" Langerhans cells because, despite the presence of the 43-kD Langerin, they do not express bona fide Birbeck granules. Thus, our results demonstrate a novel pathway in monocyte differentiation into dendritic cells.


Subject(s)
Dendritic Cells/cytology , Interleukin-15/pharmacology , Langerhans Cells/cytology , Monocytes/cytology , Monocytes/drug effects , Antigen Presentation , Cell Differentiation , Dendritic Cells/classification , Epidermal Cells , Epidermis/immunology , Granulocyte-Macrophage Colony-Stimulating Factor/pharmacology , Humans , Langerhans Cells/classification , Lymphocyte Culture Test, Mixed , Phenotype
4.
J Immunol ; 167(9): 5067-76, 2001 Nov 01.
Article in English | MEDLINE | ID: mdl-11673516

ABSTRACT

The adaptive immune system has evolved distinct responses against different pathogens, but the mechanism(s) by which a particular response is initiated is poorly understood. In this study, we investigated the type of Ag-specific CD4(+) Th and CD8(+) T cell responses elicited in vivo, in response to soluble OVA, coinjected with LPS from two different pathogens. We used Escherichia coli LPS, which signals through Toll-like receptor 4 (TLR4) and LPS from the oral pathogen Porphyromonas gingivalis, which does not appear to require TLR4 for signaling. Coinjections of E. coli LPS + OVA or P. gingivalis LPS + OVA induced similar clonal expansions of OVA-specific CD4(+) and CD8(+) T cells, but strikingly different cytokine profiles. E. coli LPS induced a Th1-like response with abundant IFN-gamma, but little or no IL-4, IL-13, and IL-5. In contrast, P. gingivalis LPS induced Th and T cell responses characterized by significant levels of IL-13, IL-5, and IL-10, but lower levels of IFN-gamma. Consistent with these results, E. coli LPS induced IL-12(p70) in the CD8alpha(+) dendritic cell (DC) subset, while P. gingivalis LPS did not. Both LPS, however, activated the two DC subsets to up-regulate costimulatory molecules and produce IL-6 and TNF-alpha. Interestingly, these LPS appeared to have differences in their ability to signal through TLR4; proliferation of splenocytes and cytokine secretion by splenocytes or DCs from TLR4-deficient C3H/HeJ mice were greatly impaired in response to E. coli LPS, but not P. gingivalis LPS. Therefore, LPS from different bacteria activate DC subsets to produce different cytokines, and induce distinct types of adaptive immunity in vivo.


Subject(s)
Drosophila Proteins , Escherichia coli/pathogenicity , Immunity/drug effects , Lipopolysaccharides/pharmacology , Porphyromonas gingivalis/pathogenicity , Animals , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , Cytokines/biosynthesis , Dendritic Cells/drug effects , Dendritic Cells/immunology , Interleukin-12/biosynthesis , Lymphocyte Activation/drug effects , Membrane Glycoproteins/physiology , Mice , Mice, Inbred C3H , Mice, Inbred C57BL , Mice, Transgenic , Ovalbumin/immunology , Receptors, Cell Surface/physiology , T-Lymphocytes, Helper-Inducer/drug effects , T-Lymphocytes, Helper-Inducer/immunology , Toll-Like Receptor 4 , Toll-Like Receptors
5.
J Exp Med ; 192(6): 769-79, 2000 Sep 18.
Article in English | MEDLINE | ID: mdl-10993908

ABSTRACT

Hyaluronan (HA), a high molecular weight glycosaminoglycan, is expressed abundantly in the extracellular matrix and on cell surfaces. Although HA is known to bind many adhesion molecules, little information has been available with respect to its direct physiological role. In this study, we developed a novel 12-mer (GAHWQFNALTVR) peptide inhibitor of HA, termed "Pep-1," by using phage display technology. Pep-1 showed specific binding to soluble, immobilized, and cell-associated forms of HA, and it inhibited leukocyte adhesion to HA substrates almost completely. Systemic, local, or topical administration of Pep-1 inhibited the expression of contact hypersensitivity responses in mice by blocking skin-directed homing of inflammatory leukocytes. Pep-1 also inhibited the sensitization phase by blocking hapten-triggered migration of Langerhans cells from the epidermis. These observations document that HA plays an essential role in "two-way" trafficking of leukocytes to and from an inflamed tissue, and thus provide technical and conceptual bases for testing the potential efficacy of HA inhibitors (e.g., Pep-1) for inflammatory disorders.


Subject(s)
Dermatitis, Contact/physiopathology , Hyaluronic Acid/physiology , Leukocytes/physiology , Oligopeptides/pharmacology , Amino Acid Sequence , Animals , Cell Adhesion/drug effects , Cell Adhesion/physiology , Cell Line , Chemotaxis, Leukocyte/drug effects , Chemotaxis, Leukocyte/physiology , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Endothelium, Vascular/physiology , Female , Hyaluronic Acid/antagonists & inhibitors , Inflammation , Keratinocytes/cytology , Keratinocytes/drug effects , Keratinocytes/physiology , Langerhans Cells/cytology , Langerhans Cells/drug effects , Langerhans Cells/physiology , Leukocytes/drug effects , Mice , Mice, Inbred BALB C , Oligopeptides/chemical synthesis , Oligopeptides/chemistry , Peptide Library , Skin/cytology , Skin/drug effects , Skin/immunology
6.
J Exp Med ; 190(1): 9-19, 1999 Jul 05.
Article in English | MEDLINE | ID: mdl-10429666

ABSTRACT

T cell recruitment to extralymphoid tissues is fundamental to the initiation and perpetuation of the inflammatory state during immune and autoimmune responses. Interleukin (IL)-15 is a proinflammatory cytokine whose described functions largely overlap with those of IL-2. The latter is attributable in large part to its binding of the heterotrimeric receptor that contains the beta and gamma chains of the IL-2R in combination with an unique IL-15R alpha chain. However, unlike IL-2, IL-15 and its receptor have a wide tissue and cell type distribution, including endothelial cells. Here, we examine the effect of IL-15 on hyaluronan expression by endothelial cells, and investigate its role in vivo in promoting the extravasation of antigen-activated T cells through a CD44-dependent pathway. The expression of hyaluronan on primary endothelial cells and microvascular endothelial cell lines is induced by IL-15, whereas IL-2 has no such activity. Moreover, intraperitoneal administration of IL-15 or TNF-alpha in the absence of other exogenous proinflammatory stimuli allows the extravasation of superantigen-stimulated T cells into this site in vivo in a CD44-dependent manner. T cell recruitment induced by IL-15 requires expression of an intact IL-2R beta chain, indicating that IL-15 operates in this context through the traditional IL-15R. The results suggest that IL-15 can regulate endothelial cell function and thereby enables a CD44-initiated adhesion pathway that facilitates entry of activated T lymphocytes into inflammatory sites. They further demonstrate a novel role for IL-15 (distinct from any of IL-2) in regulating microvascular endothelial cell adhesive function help to understand the role of IL-15R expression on endothelium, and further support a central position for this cytokine in orchestrating multiple sequential aspects of T cell effector function and therefore chronic inflammatory processes.


Subject(s)
Endothelium, Vascular/drug effects , Hyaluronic Acid/biosynthesis , Interleukin-15/pharmacology , Promoter Regions, Genetic , Animals , Cattle , Cell Line , Cell Transformation, Viral , Endothelium, Vascular/metabolism , Flow Cytometry , Humans , Hyaluronan Receptors/metabolism , Interleukin-2/pharmacology , Lymphocyte Activation , Mice , Peritoneal Cavity/cytology , Rats , Receptors, Interleukin-15 , Receptors, Interleukin-2/metabolism , Simian virus 40 , T-Lymphocytes/drug effects , T-Lymphocytes/metabolism
7.
J Clin Invest ; 101(6): 1261-72, 1998 Mar 15.
Article in English | MEDLINE | ID: mdl-9502767

ABSTRACT

The capacity of endothelial cells (EC) to produce IL-15 and the capacity of IL-15 to influence transendothelial migration of T cells was examined. Human umbilical vein endothelial cells expressed both IL-15 mRNA and protein. Moreover, endothelial-derived IL-15 enhanced transendothelial migration of T cells as evidenced by the inhibition of this process by blocking monoclonal antibodies to IL-15. IL-15 enhanced transendothelial migration of T cells by activating the binding capacity of the integrin adhesion molecule LFA-1 (CD11a/CD18) and also increased T cell motility. In addition, IL-15 induced expression of the early activation molecule CD69. The importance of IL-15 in regulating migration of T cells in vivo was documented by its capacity to enhance accumulation of adoptively transferred human T cells in rheumatoid arthritis synovial tissue engrafted into immune deficient SCID mice. These results demonstrate that EC produce IL-15 and imply that endothelial IL-15 plays a critical role in stimulation of T cells to extravasate into inflammatory tissue.


Subject(s)
Arthritis, Rheumatoid/immunology , Endothelium, Vascular/immunology , Endothelium, Vascular/metabolism , Interleukin-15/immunology , Interleukin-15/metabolism , T-Lymphocytes/immunology , Adoptive Transfer , Animals , Antibodies, Blocking/immunology , Antibodies, Monoclonal/immunology , Antigens, CD/metabolism , Antigens, Differentiation, T-Lymphocyte/metabolism , Arthritis, Rheumatoid/metabolism , Cell Adhesion/immunology , Cell Movement/immunology , Cells, Cultured , Flow Cytometry , Fluorescent Antibody Technique, Indirect , Humans , Immunohistochemistry , Inflammation/immunology , Inflammation/metabolism , Intercellular Adhesion Molecule-1/metabolism , Lectins, C-Type , Lymphocyte Function-Associated Antigen-1/metabolism , Mice , Mice, SCID , RNA, Messenger/metabolism , Synovial Membrane/immunology , Synovial Membrane/metabolism , Synovial Membrane/transplantation , T-Lymphocytes/cytology , T-Lymphocytes/metabolism , Tissue Transplantation , Umbilical Veins/cytology , Umbilical Veins/immunology
8.
J Clin Invest ; 101(1): 97-108, 1998 Jan 01.
Article in English | MEDLINE | ID: mdl-9421471

ABSTRACT

The localization of circulating leukocytes within inflamed tissues occurs as the result of interactions with and migration across vascular endothelium, and is governed, in part, by the expression of adhesion molecules on both cell types. Recently, we have described a novel primary adhesion interaction between the structurally activated form of the adhesion molecule CD44 on lymphocytes and its major ligand hyaluronan on endothelial cells under physiologic laminar flow conditions, and have proposed that this interaction functions in an extravasation pathway for lymphocytes in vascular beds at sites of inflammation. While the regulation of activated CD44 on leukocytes has been characterized in depth, regulation of hyaluronate (HA) on endothelial cells has not been extensively studied. Here we demonstrate that the expression of HA on cultured endothelial cell lines and primary endothelial cultures is inducible by the proinflammatory cytokines TNFalpha and IL-1beta, as well as bacterial lipopolysaccharide. In addition, this inducibility appears strikingly restricted to endothelial cells derived from microvascular, but not large vessel, sources. The elevated HA levels thus induced result in increased CD44-dependent adhesive interactions in both nonstatic shear and laminar flow adhesion assays. Changes in mRNA levels for the described HA synthetic and degradative enzymes were not found, suggesting other more complex mechanisms of regulation. Together, these data add to the selectin and immunoglobulin gene families a new inducible endothelial adhesive molecule, hyaluronan, and help to further our understanding of the potential physiologic roles of the CD44/HA interaction; i.e., local cytokine production within inflamed vascular beds may enhance surface hyaluronan expression on endothelial cells, thereby creating local sites receptive to the CD44/HA interaction and thus extravasation of inflammatory cells.


Subject(s)
Endothelium, Vascular/metabolism , Glycosyltransferases , Hyaluronan Receptors/metabolism , Hyaluronic Acid/biosynthesis , Inflammation/metabolism , Membrane Proteins , Transferases , Xenopus Proteins , Animals , Cattle , Cell Adhesion , Cell Line , Cell Line, Transformed , Cytokines/pharmacology , Endothelium, Vascular/cytology , Endothelium, Vascular/drug effects , Glucuronidase/genetics , Glucuronidase/metabolism , Glucuronosyltransferase/genetics , Glucuronosyltransferase/metabolism , Humans , Hyaluronan Synthases , Hyaluronoglucosaminidase/genetics , Hyaluronoglucosaminidase/metabolism , Inflammation/pathology , Interferon-gamma/pharmacology , Interleukin-1/pharmacology , Interleukin-12/pharmacology , Lipopolysaccharides/pharmacology , Lymph Nodes/cytology , Mice , RNA, Messenger/metabolism , Tumor Necrosis Factor-alpha/pharmacology , beta-N-Acetylhexosaminidases/genetics , beta-N-Acetylhexosaminidases/metabolism
9.
Arch Dermatol Res ; 289(8): 435-9, 1997 Jul.
Article in English | MEDLINE | ID: mdl-9266019

ABSTRACT

We took advantage of the recently generated 4F7 mAb, which recognizes an epitope expressed on dendritic cells (DC) from different tissues, to freshly isolate and positively sort for these cells and to characterize their cytokine pattern and antigen-presenting capacity in comparison with epidermal Langerhans cells (LC). RT-PCR and Northern blot analyses demonstrated constitutive mRNA expression of MIP-1 gamma, MIP-1 alpha, C10, and IL-1 beta in both 4F7+ DC and LC. Lipopolysaccharide (LPS) treatment resulted in the upregulation of mRNA expression of all four cytokines and in a newly detected signal for TNF alpha. Immunoblot analysis showed constitutive secretion of MIP-1 gamma, with LPS treatment resulting in the upregulation of IL-1 beta production and in newly detected TNF alpha secretion. 4F7+ DC were also shown to express mRNA for the common gamma chain receptor of IL-2 and for the receptor of IL-4. Finally, we demonstrated freshly isolated 4F7+ DC to be equivalent to freshly isolated LC in their capacity to present alloantigen in the mixed leukocyte reaction (MLR) and to process and present purified protein derivative (PPD) to Th1 and Th2 clones. We conclude that 4F7 is a useful marker for positively sorting DC from dermis, spleen, and lymph nodes. Regardless of tissue source, 4F7+ DC exhibit uniform cytokine and antigen-presenting capacity profiles that mimic the properties of freshly isolated epidermal LC.


Subject(s)
Antigen Presentation , Cytokines/biosynthesis , Dendritic Cells/physiology , Lymph Nodes/immunology , Skin/immunology , Spleen/immunology , Animals , Cytokines/genetics , Female , Mice , Mice, Inbred BALB C , Mice, Inbred CBA , RNA, Messenger/analysis
10.
J Invest Dermatol ; 107(5): 738-42, 1996 Nov.
Article in English | MEDLINE | ID: mdl-8875959

ABSTRACT

Granzymes, a family of serine proteases contained in cytoplasmic granules of cytotoxic T lymphocytes and natural killer cells, play a critical role in killing tumor targets by triggering rapid breakdown of DNA and subsequent apoptosis. We have reported previously that dendritic epidermal T cells, which are skin-specific members of the tissue-type gamma(delta) T-cell family in mice, are capable of killing selected tumor cell lines. Here we report that short-term cultured dendritic epidermal T-cell lines contain significant N-alpha-benzyloxycarbonyl-L-Lys-thiobenzyl esterase activity, produce granzyme A protein, and express constitutively mRNA for granzymes A and B. Messenger RNA expression for granzyme B was also confirmed in freshly procured Thy-1+ epidermal cells (i.e., dendritic epidermal T cells). Finally, preincubation of dendritic epidermal T cell lines with a granzyme inhibitor, dichloroisocoumarin, but not with a cysteine protease inhibitor, E-64, abrogated completely their capacity to trigger DNA breakdown in YAC-1 target cells. These results reinforce the concept that dendritic epidermal T cells represent skin-resident killer cells that share several functional properties with conventional killer leukocytes, thereby playing a local immunosurveillance role against tumor development.


Subject(s)
Cytotoxicity, Immunologic , Receptors, Antigen, T-Cell, gamma-delta/analysis , Serine Endopeptidases/physiology , T-Lymphocytes/immunology , Animals , DNA Fragmentation , Dendritic Cells/physiology , Female , Granzymes , Mice , Mice, Inbred CBA
11.
J Invest Dermatol ; 107(4): 582-8, 1996 Oct.
Article in English | MEDLINE | ID: mdl-8823364

ABSTRACT

Exposure to certain allergens via epithelial tissues is the primary route for the induction of immunoglobulin E-dependent allergies of the immediate type associated with atopic diseases. In order to address the question whether and how epithelial cells might contribute to the induction or increase of TH2-dependent IgE production, we performed co-culture experiments of syngeneic epidermal cells and cells from the associated lymphoid tissue or spleen (responder cells) of BALB/c mice primed with ovalbumin in vivo. In the presence of ovalbumin in vitro, immunoglobulin E but not immunoglobulin G2a production was significantly enhanced by the addition of epidermal cells, and separation of epidermal cells from responder cells by a membrane that prevented cellular contacts or addition of antibodies against intercellular adhesion molecule-1 reduced the enhancement of immunoglobulin E production induced by epidermal cells. Depletion of major histocompatibility complex class II+ antigen presenting Langerhans cells from the epidermal cells prior to co-culture also reduced the enhancement of immunoglobulin E production induced by epidermal cells. The enhanced immunoglobulin E production was dependent on the induction of TH2 cell-derived interleukin-4 detected in co-cultures because it was completely inhibited after addition of anti-interleukin-4 antibodies that also lead to increased immunoglobulin G2a production. Whereas interleukin-4 was not produced by epidermal cells, interleukin-10 seemed to be one important mediator contributed by epidermal cells. Interleukin-10 skewed the response toward a TH2-mediated IgE response because antibodies against interleukin-10 inhibited interleukin-4 and immunoglobulin E production, whereas they enhanced interferon-gamma and immunoglobulin G2a production.


Subject(s)
Allergens/immunology , Epidermis/immunology , Epidermis/metabolism , Immunoglobulin E/biosynthesis , Interleukin-4/biosynthesis , Ovalbumin/immunology , Animals , Dendritic Cells/metabolism , Dose-Response Relationship, Drug , Epidermal Cells , Female , Histocompatibility Antigens Class II/analysis , Immunization , Interleukin-10/genetics , Interleukin-10/physiology , Interleukin-4/physiology , Keratinocytes/physiology , Langerhans Cells/immunology , Lymphoid Tissue/cytology , Lymphoid Tissue/metabolism , Mice , Mice, Inbred BALB C , RNA, Messenger/biosynthesis
12.
J Immunol ; 156(9): 3102-6, 1996 May 01.
Article in English | MEDLINE | ID: mdl-8617929

ABSTRACT

Langerhans cells (LC) are skin-specific members of the dendritic cell (DC) family. DC are unique among APC for their capacity to activate immunologically naive T cells, but little is known about their chemotactic recruitment of T cells. We now report that LC produce macrophage inflammatory protein-1 gamma (MIP-1 gamma), a newly identified CC chemokine. MIP-1 gamma mRNA was detected in epidermal cells freshly procured from BALB/c mice, and depletion of I-A+ epidermal cells (i.e., LC) abrogated that expression. MIP-1 gamma mRNA was detected in the XS52 LC-like DC line as well as by 4F7+ splenic DC and granulocyte-macrophage CSF-propagated bone marrow DC. XS52 DC culture supernatants contained 9 and 10.5 kDa immunoreactivities with anti-MIP-1 gamma Abs. We observed in Boyden chamber assays that 1) XS52 DC supernatant (added to the lower chambers) induced significant migration by splenic T cells; 2) this migration was blocked by the addition of anti-MIP-1 gamma in the lower chambers or by rMIP-1 gamma in the upper chambers; and 3) comparable migration occurred in both CD4+ and CD8+ T cells and in both activated and nonactivated T cells. We conclude that mouse DC (including LC) have the capacity to elaborate the novel CC chemokine MIP-1 gamma, suggesting the active participation of DC in recruiting T cells before activation.


Subject(s)
Chemokines/biosynthesis , Dendritic Cells/metabolism , Monokines/biosynthesis , Animals , Cell Line , Chemokine CCL4 , Chemokines/genetics , Chemokines/physiology , Chemotaxis, Leukocyte/drug effects , Female , Macrophage Inflammatory Proteins , Mice , Mice, Inbred BALB C , Monokines/genetics , Monokines/physiology , RNA, Messenger/biosynthesis , T-Lymphocytes/physiology
13.
Photodermatol Photoimmunol Photomed ; 12(1): 17-21, 1996 Feb.
Article in English | MEDLINE | ID: mdl-8884894

ABSTRACT

The purposes of the present study were to determine whether endothelial cells express IL-15 and to evaluate effects of ultraviolet B (UVB) and 8-methoxypsoralens plus UVA (PUVA) on such expression. Cultured human endothelial cells derived from dermis or umbilical veins were subjected to reverse transcriptase-polymerase chain reaction (RT-PCR) and immunoblot analyses for the detection of IL-15 mRNA and protein, respectively. Both dermal and umbilical vein endothelial cells were shown to express IL-15 mRNA and protein, and these markers were upregulated following UVB or PUVA treatment (but not by UVA or 8-methoxypsoralens alone). Also using RT-PCR, dermal and umbilical vein endothelial cells were shown to express IL-2R gamma c mRNA. These results expand the sources of IL-15 in skin to include keratinocytes, dermal fibroblasts, and now endothelial cells. That IL-15 from all three skin cells can be upregulated by UV treatment suggests a role for this cytokine in photosensitive disorders. Finally, the possibility of an autocrine effect of IL-15 on endothelial cells is raised by the expression of IL-2R gamma c in these cells.


Subject(s)
Endothelium, Vascular/metabolism , Ficusin/pharmacology , Interleukin-15/biosynthesis , Photosensitizing Agents/pharmacology , Ultraviolet Rays , Up-Regulation/radiation effects , Cells, Cultured , Endothelium, Vascular/drug effects , Endothelium, Vascular/radiation effects , Humans , Immunoblotting , PUVA Therapy , Polymerase Chain Reaction , Skin/blood supply , Umbilical Veins/cytology , Up-Regulation/drug effects
14.
Eur J Immunol ; 26(1): 156-60, 1996 Jan.
Article in English | MEDLINE | ID: mdl-8566059

ABSTRACT

The common cytokine receptor gamma chain (gamma c) is an indispensable component of interleukin (IL)-2, IL-4, IL-7, IL-9, and IL-15 receptors, and its expression has been detected in several leukocyte populations, including T cells, B cells, monocytes, natural killer cells, and neutrophils. The purpose of this study was to determine whether gamma c receptors are expressed by dendritic cells (DC). Constitutive gamma c mRNA expression was observed by reverse transcription polymerase chain reaction and/or Northern blotting for: (a) Ia+ epidermal Langerhans cells (LC), (b) 4F7+ splenic DC, (c) granulocyte/macrophage colony-stimulated factor-propagated bone marrow-derived DC, and (d) the epidermal-derived DC line, XS52, which retains important functions of epidermal LC. Exposure of XS52 cells to recombinant IL-4 induced a rapid up-regulation of c-myc mRNA expression, and this IL-4-dependent signaling was blocked almost completely by anti-gamma c monoclonal antibody (mAb) TUGm2 in a soluble form. Moreover, c-myc up-regulation was inducible in XS52 cells by the same mAb in an immobilized form. These results imply that molecules recognized by this antibody (i.e. gamma c receptors) are expressed on XS52 cell surfaces. We thus conclude that DC express functional gamma c receptors, which then mediate cytokine-dependent regulation of DC functions.


Subject(s)
Dendritic Cells/metabolism , Epidermis/immunology , Langerhans Cells/metabolism , Receptors, Cytokine/metabolism , Animals , Base Sequence , Cell Line , Dendritic Cells/classification , Female , Membrane Proteins/biosynthesis , Mice , Mice, Inbred BALB C , Molecular Sequence Data , RNA, Messenger/biosynthesis , Receptors, Cytokine/analysis , Receptors, Cytokine/genetics
15.
J Invest Dermatol ; 105(6): 837-43, 1995 Dec.
Article in English | MEDLINE | ID: mdl-7490480

ABSTRACT

Dendritic epidermal T cells (DETC) are skin-specific members of the epithelial gamma delta T-cell family in mice. We have reported previously that the growth of DETC is promoted by interleukin (IL)-2 in an autocrine fashion, or by IL-7, which is secreted by neighboring keratinocytes. Here we report that DETC growth is promoted by IL-15, a newly discovered T-cell growth factor that is produced in lymphoid as well as nonlymphoid tissues. Recombinant IL-15 promoted the growth of the 7-17 DETC line in a time- and dose-dependent fashion. Using monoclonal antibodies against alpha-, beta-, or gamma c-chains of the IL-2 receptor complex, we observed that the combination of anti-beta chain and anti-gamma c chain antibodies blocked IL-15 responsiveness completely, whereas anti-alpha chain had no effect. These results indicate that this gamma delta T-cell line uses the beta/gamma c heterodimer for proliferative responses to IL-15. Antibodies against IL-2 or IL-7 did not block IL-15-driven proliferation of 7-17 DETC, indicating that IL-15 promotes their growth in an IL-2- and IL-7-independent manner. Both the surface expression of beta/gamma c heterodimers and the IL-15 responsiveness of 7-17 DETC were highest 1 to 8 days after concanavalin A stimulation, and both declined substantially 21 days after stimulation, illustrating regulation by the state of cell activation. Working with epidermal cells that were freshly procured from CBA mice, we noted that IL-15 promoted conavalin-A-triggered growth of Thy-1+ cells (i.e., DETC), but not of the Thy-1- cells. The gamma c-chain was not expressed by freshly procured DETC, becoming detectable within 48 h after concanavalin A stimulation. We propose that IL-15 facilitates the growth of epithelial gamma delta T cells by a beta/gamma c receptor-dependent mechanism.


Subject(s)
Dendritic Cells/drug effects , Epidermal Cells , Interleukins/pharmacology , Lymphocyte Activation/drug effects , Receptors, Antigen, T-Cell, gamma-delta/analysis , Receptors, Interleukin-2/physiology , T-Lymphocytes/drug effects , Animals , Cell Line , Female , Humans , Interleukin-15 , Mice , Mice, Inbred AKR , Mice, Inbred CBA , T-Lymphocytes/immunology
16.
J Immunol ; 155(9): 4492-6, 1995 Nov 01.
Article in English | MEDLINE | ID: mdl-7594612

ABSTRACT

Ultraviolet B (UVB) radiation is a potent modulator of skin-related immune responses, particularly those involving the synthesis and the secretion of cytokines. The discovery of a new T cell mitogen, IL-15, prompted use to investigate its expression in skin and to examine the effects of UVB radiation on such expression. RNA from unirradiated and UVB-irradiated epidermal and dermal sheets derived from human foreskin as well as from unirradiated and UVB-irradiated skin cell populations were assayed for IL-15 expression by semiquantitative RT-PCR. Constitutive levels of IL-15 mRNA were detected in dermal sheets, but not in epidermal sheets. Following UVB treatment, IL-15 mRNA was induced in epidermal sheets and enhanced in dermal sheets. UVB-inducible epidermal expression of IL-15 mRNA was traced to HLA-DR- cells (presumably keratinocytes) and not to HLA-DR+ cells (Langerhans cells). Cultured keratinocytes and dermal fibroblasts displayed basal levels of IL-15 mRNA that were also up-regulated following UVB exposure. Immunoblot analysis revealed secretion of IL-15 protein by keratinocytes that was enhanced following UVB treatment. These results constitute the first report of IL-15 mRNA expression and protein production in human skin. In addition to expanding the known influence of UVB radiation on the capacity of keratinocytes and dermal fibroblasts to express immunomodulatory cytokines, these findings suggest a new mechanism by which UVB can promote Ag-independent T cell responses via elaboration of IL-15.


Subject(s)
Interleukins/radiation effects , Skin/radiation effects , Ultraviolet Rays , Up-Regulation/radiation effects , Base Sequence , Cells, Cultured , Dose-Response Relationship, Radiation , Fibroblasts/radiation effects , Humans , Interleukin-15 , Interleukins/genetics , Interleukins/metabolism , Keratinocytes/metabolism , Keratinocytes/radiation effects , Male , Molecular Sequence Data , RNA, Messenger/radiation effects , Skin/cytology , Up-Regulation/genetics
17.
Exp Dermatol ; 4(5): 291-6, 1995 Oct.
Article in English | MEDLINE | ID: mdl-8589919

ABSTRACT

Heat-stable antigen (HSA), expressed by various antigen-presenting cells (APC), has been described as a costimulatory molecule for CD4+ T cells. Recently, we observed that HSA also serves as an important costimulatory molecule on epidermal Langerhans cells (LC). During these studies, low levels of HSA staining were also detected on normal murine keratinocytes (KC). To investigate whether HSA also is involved in T-cell activation by KC, normal murine KC or the spontaneously transformed KC cell-line PAM 212 were treated with PDB or PMA to induce HSA-expression. FACS analyses showed induction of HSA expression on normal murine KC, as well as PAM 212 cells. In functional assays PDB or PMA-treated normal or transformed KC were far more potent inducers of primary allogeneic T-cell responses than untreated KC. Addition of anti-HSA-specific mAb 20C9 specifically inhibited the costimulatory activity of KC, an effect that was even more pronounced when CTLA-4Ig was added to the cultures. Cleavage of HSA on KC surfaces by a phosphoinositol-specific phospholipase C (PI-PLC) also significantly inhibited the costimulatory capacity of KC for naive CD4+ T cells. In aggregate, our data indicate that expression of HSA on activated KC contributes to the capacity of these cells to induce proliferation of allogeneic T cells.


Subject(s)
Antigens, CD/immunology , Immunization , Keratinocytes/immunology , Lymphocyte Activation , T-Lymphocytes/physiology , Animals , Antibodies, Monoclonal/pharmacology , Antigens, CD/genetics , Base Sequence , Cells, Cultured , Keratinocytes/drug effects , Keratinocytes/physiology , Lymph Nodes/cytology , Mice , Mice, Inbred BALB C , Mice, Inbred C3H , Molecular Probes/genetics , Molecular Sequence Data , Phorbol Esters/pharmacology , Phosphatidylinositol Diacylglycerol-Lyase , Phosphoinositide Phospholipase C , Phosphoric Diester Hydrolases/pharmacology , RNA, Messenger/metabolism , T-Lymphocytes/drug effects
18.
J Invest Dermatol ; 105(2): 220-4, 1995 Aug.
Article in English | MEDLINE | ID: mdl-7636304

ABSTRACT

Superantigens are potent modulators of the immune system, especially T cells. Therefore, we determined the influence of superantigens on the T-cell-mediated immune response, contact sensitivity. We chose the combination of staphylococcal enterotoxin B (SEB) as superantigen and 2,4-dinitrofluorbenzene (DNFB) as the contact sensitizer, because in BALB/c mice SEB reacts almost exclusively with V beta 8+ T cells, and these cells are capable of transferring contact sensitivity to DNFB from sensitized donors to naive syngeneic recipients. Pretreatment with a single intradermal injection of 50 ng SEB 24 h before DNFB exposure at the same site on the lower abdomen enhanced the induction of contact sensitivity: its intradermal injection permitted sensitization with non-sensitizing concentrations of DNFB as assessed by ear swelling responses after challenge with DNFB. In contrast, pretreatment with repeated intradermal injections of 50 ng SEB every other day over at least 1 week inhibited the induction of contact sensitivity following sensitization. The enhancing effect of SEB may be explained by the creation of a proinflammatory milieu in the skin after a single intradermal injection of the bacterial toxin, whereas the inhibitory effect may be due to tolerization of V beta 8+ T cells. The data indicate that products of skin-colonizing bacteria that can serve as superantigens are able to augment or inhibit the development of contact sensitivity.


Subject(s)
Antigens, Bacterial/immunology , Dermatitis, Contact/immunology , Superantigens/immunology , Animals , Dinitrofluorobenzene/immunology , Down-Regulation , Enterotoxins/pharmacology , Female , Immunization , Interferon-gamma/biosynthesis , Lymphoid Tissue/metabolism , Mice , Mice, Inbred BALB C , Receptors, Antigen, T-Cell, alpha-beta/metabolism , Skin , T-Lymphocytes/immunology , T-Lymphocytes/metabolism
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