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CHEMOKINES
From the Departments of Biochemistry/Molecular Biology
and Hematology/ Oncology, Indiana University Cancer Center, Indiana
University School of Medicine, Indianapolis; and Joint Program in
Transfusion Medicine, Children's Hospital, and Department of
Pathology, Harvard Medical School, Boston, MA.
Chemokines are a large family of cytokines that direct normal
leukocyte migration. They also have been implicated in leukocyte development and in the pathogenesis of many diseases. The CC chemokine CCL21, also known as Exodus-2, SLC, 6Ckine, and TCA4 induces both the
adhesion and migration of human T cells. CCL21 is hypothesized to
regulate the trafficking of T cells through secondary lymphoid tissues.
To test this hypothesis, a transgenic mouse model was generated that
placed the expression of mouse CCL21 (mCCL21) under the control of the
T cell-specific lck promoter to abrogate the concentration gradient to
which T cells normally respond. Overexpression of mCCL21 in T cells
resulted in defects in CCL21- and CCL19-induced T-cell chemotaxis, node
T-cell subpopulations, and lymph node architecture. The regulation of
T-cell trafficking in secondary lymphoid tissues by CCL21 is therefore
a tightly regulated system that can be altered by changes in the level
of environmental CCL21 protein.
(Blood. 2001;98:3562-3568) Chemokines are a large family of cytokines that
direct leukocyte migration and have been implicated in the regulation
of leukocyte development, angiogenesis, tumor growth, and
metastasis.1-4 They play a key role in both innate and
acquired immunity.5-8 The number and spacing of the first
cysteines in the amino acid sequence are used to characterize
chemokines into 4 subfamilies: XCL1-2, CCL1-27, CXCL1-14, and
CX3CL1.9-10 Chemokines act through chemokine receptors (XCR1, CCR1-11, CXCR1-5, CX3CR1), which are a
subfamily of G protein-coupled 7-transmembrane
receptors.9-11
Chemokines are reported to be mediators of tissue destruction in a wide
variety of human diseases, including autoimmune disease, postinfarct
reperfusion injury, atherosclerosis, adult respiratory distress
syndrome, and infectious disease.1-8,12-17 For example, CCL11, CCL7, and CCL5 are all suspected mediators of asthma due to
their localized presence at the disease site and function in attracting
eosinophils.5 Chemokines also stimulate neutrophil infiltration of infarcted tissue, mediating reperfusion tissue damage.18 In addition, the human immunodeficiency virus
(HIV) coreceptors that function with CD4 are chemokine receptors,
suggesting a mechanism for the inhibition of HIV infection by specific
chemokines.19-21
Chemokines have been implicated in angiogenesis, tumor growth, and
metastasis.3,5,8 It has been reported that ELR-CXC chemokines are angiogenic and most non-ELR-CXC chemokines are angiostatic.3 In addition, the chemokine receptor CXCR4
mediates metastasis in breast cancer.22 Chemokines have
generated interest as tumor vaccines for their chemoattractant and
T-cell and natural killer (NK)-cell stimulatory properties. The CC
chemokine CCL1 can induce tumor-specific and long-lasting immunity in
mice that have been immunized with tumor cells expressing
CCL1.23 Tumor cells transduced with the CC chemokine CCL2
also stimulate a specific immune response that protects immunized rats
against further tumor cell challenge.24 XCL1 transduced
into cells, simultaneously with IL-2 as a T cell-stimulatory molecule,
was reported to be effective as a tumor vaccine.25
CCL1926,27 has been shown to inhibit breast cancer cell
growth in a murine model by an NK and CD4+ cell-mediated
mechanism.28 We recently found that CCL19, 20, and 21 inhibited chronic myelogenous leukemia progenitors.29 CCL21 has also been reported to stimulate lymphocyte antitumor responses in mouse tumor models and cell line-based
models.30-32
Our laboratory and others cloned CCL21, thought to be a critical
regulator of T-cell trafficking through lymphoid
tissue.33-36 We and others found that CCL21 expression in
the high endothelial venule (HEV) of the lymph node stimulates the
adherence and migration of T cells into the secondary lymphoid tissue
where antigen presentation can occur.37-41 CCL21
expression attracts and colocalizes both T cells and antigen-presenting
dendritic cells to the nodal T-cell zone where antigen presentation can
occur.42-46 The importance of CCL21 and CCL19 in T-cell
trafficking has also been implied by studies involving
plt-mice and CCR7-null mice. The plt (paucity of
lymph node T cells) mouse lacks CCL19 and CCL21 expression and displays
an inability of T cells and activated dendritic cells to traffic to
lymph nodes or T-cell zones of the spleen.47-53 CCL19 and
CCL21 both bind CCR7.40,42-46 Data analysis of
CCR7-deficient mice shows the disordered migration of lymphocytes to
secondary lymphoid organs and a failure of skin dendritic cells to
migrate into draining lymph nodes.54
In this study, a transgenic mouse overexpressing CCL21 in T cells
was generated to abrogate the CCL21 concentration gradient T cells
encounter and migrate toward. We found that overexpression of CCL21 by
T cells in the CCL21 transgenic mice resulted in defects in mCCL21- and
mCCL19-induced T-cell chemotaxis, lymphocyte subpopulations in lymphoid
tissues, and nodal architecture.
Generation of the lck-mCCL21 construct
Microinjection and screening of lck-mCCL21 construct
Protein analysis
Chemotaxis Chemotaxis assays were performed using 96-well chemotaxis chambers (Neuroprobe, Gaithersburg, MD) in accordance with manufacturer's instructions with minor variations as described.58-61 Briefly, 0, 200, 400, and 800 ng/mL of the appropriate murine chemokine was added to 300µL RPMI without phenol media supplemented with 10% fetal bovine serum (FBS) in the lower chamber. Two hundred thousand fluorescent-tagged (4 µg/mL Calcein AM, Molecular Probes, Eugene, OR) isolated lymphocytes or splenocytes in 50 µL media were added to the upper side of the membrane (5.7-mm diameter, 5-µm pore size, polycarbonate membrane).Total cell migration was obtained by measuring fluorescence (excitation, 485 nm; emission, 530 nm) and calculating cell number in the lower well by comparison to a cell number standard curve after 3 hours of incubation at 37°C, 5% CO2. Percent migration was calculated by dividing the number of the cells in the lower well by the total cell input multiplied by 100 and subtracting random migration to the lower chamber without chemokine presence. Cell subtype migration was obtained by staining the input cell population and migrating cell population with fluorochrome-conjugated monoclonal antibodies to CD3, CD19, CD4, CD8, and 62L as described below. The numbers of each cell type in the input and migrating populations were then used to obtain the percent migration of each cell type. This was done by dividing the absolute number of cells of a given cell type in the lower well by the total input cell number of that same cell type, multiplied by 100, and then subtracting random transwell migration to the lower chamber without chemokine presence. At least 4 WT and transgenic individual mice were analyzed separately in triplicate, and then the data averaged for statistical analysis. Adhesion Static adhesion assays were performed as we previously described.37,43 Briefly, intercellular adhesion molecule 1 (ICAM-1) was coated on glass slides and lymphocytes were allowed to settle for 10 minutes on the surface of the slide. CCL21 was added and nonadherent cells were washed away. Adherent cells were then fixed and counted. Adhesion assays were performed on splenocytes from lck-mCCL21 transgenic mice and compared to normal control mouse splenocytes.Flow cytometry Cells were stained with fluorochrome-conjugated monoclonal antibodies to CD3, CD19, CD4, CD8, and 62L (BD Pharmingen, San Diego, CA) in accordance with the manufacturer's specifications and then counted by flow cytometric analysis. CD3/62L was used to identify lymph node-homing murine T cells. Harvested cells are washed in phosphate-buffered saline (PBS)/penicillin/streptomycin/1% bovine serum albumin (BSA) and resuspended in 100 µL PBS/BSA. Then 0.5 µg of the appropriate antibody was added and mixed, and the cells were incubated at 4°C in the dark for 30 minutes. The cells were then washed twice in PBS/BSA and fixed in PBS/1% paraformaldehyde for flow cytometric analysis at 488 nm. Ten thousand events were accumulated for each analysis. At least 4 WT and transgenic individual mice were analyzed separately in triplicate and the data averaged for statistical analysis.Immunohistochemistry Comparisons of T-cell and B-cell localization between normal and transgenic mice within lymphoid tissue architecture were made using immunohistology as we described.62 Freshly isolated adult mouse spleen, thymus, and lymph node tissues were frozen and cut into tissue sections. Sections were stained with biotinylated CD3 and CD19 cell surface marker antibodies (BD Pharmingen). Slides were stained with antibody and signals were developed by peroxidase/3,3'-diamobenzidine (DAB) staining in accordance with manufacturer's specifications (Vector Labs, Burlingame, CA). Slides were counterstained with hematoxylin nuclear counterstain (Vector Labs, Burlingame, CA).
DNA and protein analysis of transgenic expression of CCL21 Homozygous transgenic (lck/lck-mCCL21) and heterozygous transgenic (WT/lck-mCCL21) mice were compared to wild-type (WT/WT) mice. A Southern blot analysis of isolated mouse genomic DNA using a radiolabeled mCCL21 cDNA probe revealed the presence of the lck-mCCL21 construct in the heterozygous WT/lck-mCCL21 mice. The lck/lck-mCCL21 homozygous transgenic mice had 2 copies of the transgene (Figure 2).
The well-characterized T cell-specific lck promoter has been shown by lck-green fluorescent protein (GFP) expression to be expressed in the earliest thymic T-cell population and in splenic T cells.56 Western analysis of isolated mouse thymocyte cell lysates showed overexpression of the CCL21 protein by the lck-mCCL21 construct (Figure 2). Endogenous CCL21 protein was undetectable in the normal mouse thymocytes by Western blot analysis at this exposure, but can be seen on prolonged exposure. A copy number-dependent increase in protein expression can be seen when comparing heterozygous (WT/lck-mCCL21) and homozygous (lck/lck-mCCL21) transgenic expression of CCL21. Two mouse lineages were obtained for study that resulted from 2 distinct integration events. Both lines showed transmission of the transgene and transgenic overexpression of CCL21 protein at equal levels. Homozygous mice from both lineages were used for phenotypic analysis to ensure that observations are the direct result of CCL21 overexpression and were not dependent on the integration site. Chemotaxis To analyze total T-cell migratory function, chemotaxis assays were performed comparing WT and homozygous transgenic CD3+ mouse splenocytes and thymocytes. Flow cytometric analysis was performed to characterize starting and migrating T-cell populations. This analysis found significant defects in transgenic CD3+ splenocyte and thymocyte chemotaxis in response to mCCL21 and mCCL19 but not CXCL12 (Figure 3). Transgenic spleen T cells had a maximum decrease of 57% of their CCL21-induced chemotactic ability as compared to normal spleen T cells (P = .0004). In a similar manner, transgenic spleen T cells had a maximum decrease of 37% of their CCL19-induced chemotactic ability as compared to normal spleen T cells (P = .0014). However, no appreciable difference was seen between the ability of transgenic versus normal spleen CD3+ cells to migrate in response to CXCL12 (Figure 3, P = .445). Transgenic thymic CD3+ cells also had defects in CCL19- and CCL21-induced chemotaxis as compared to WT controls (Figure 3). Transgenic thymic T cells had maximum decreases of 60% (P = .0055) and 33% (P = .012) compared to WT/WT for CCL21- and CCL19-induced chemotaxis, respectively. Thymic T cells expressed the receptor for CXCL12 poorly and, as a result, there was little thymic T-cell chemotaxis with which to compare transgenic to WT mice.
CCL21 has been reported to mediate T-cell homing to lymph nodes via
stimulation of HEV adhesion and then chemotaxis into specific regions
within the node.42-45 To analyze lymph node- homing
T-cell migratory function, chemotaxis assays were performed comparing normal and homozygous transgenic CD3+/CD62L+
splenocytes and thymocytes (Figure 4).
CD62L expression on T cells is an absolute requirement for HEV lymph
node homing. Significant deficiencies in transgenic
CD3+/CD62L+ splenocyte and thymocyte chemotaxis
in response to CCL21 and CCL19 were observed. There was a 61%
decrease in the chemotactic response to CCL21 of
CD3+/CD62L+ splenocytes in transgenic mice as
compared to normal controls (P = .001, Figure 4).
Chemotaxis to CCL19 was also reduced by 49% in transgenic
CD3+/CD62L+ splenocytes as compared to normal
controls (P = .001, Figure 4). Chemotactic response to
CXCL12 was unchanged (Figure 4).
Mouse thymocyte CD3+/CD62L+ chemotactic response in homozygous transgenic mice was also reduced. CD3+/CD62L+ thymocytes from transgenic mice had a 62% loss in chemotactic response to 400 ng/mL CCL21 compared to normal controls (P = .003, Figure 4). Migration toward 400 ng/mL CCL19 was reduced 34% in transgenic compared to WT mice (P = .001, Figure 4). Adhesion Adhesion studies using mCXCL12, mCCL21, and mCCL19 show no significant difference between the adhesion of normal (WT/WT) or transgenic (lck/lck-mCCL21) splenocytes or thymocytes to capillary tubes coated with ICAM-1 (Figure 5).
Lymphocyte populations Cells were stained with fluorochrome-conjugated monoclonal antibodies (mAbs) to CD3, CD19, CD62L, CD4, and CD8 and then counted using flow cytometry. Analysis of CD3+ versus CD19+ (Figure 6A) showed no difference in the relative percent of T cells versus B cells in the spleen, thymus, or lymph nodes. There were no size differences between WT and transgenic nodes.
Analysis of CD4+, CD8+, and
CD4+/CD8+ double-positive cells (Figure 6B)
showed a slight increase (P = .012) in
CD4+/CD8+ double-positive T cells in the thymus
of the transgenic mice (77.8% ± 0.4%) when compared to normal mice
(WT/WT) (73.5% ± 0.3%). Analysis of
CD3+/CD62L+ versus
CD3+/CD62L Lymphoid architecture Immunohistochemistry of WT/WT and lck/lck-mCCL21 transgenic mice revealed that mCCL21 transgenic mice had abnormal lymph node architecture (Figure 7). Normal mouse and transgenic mouse lymph node frozen sections were stained for T cells (CD3) and B cells (CD19). Normal nodal architecture contained a large T-cell zone surrounded by a typical B-cell follicle. Overexpression of mCCL21 specifically in T cells results in a much smaller T-cell zone and a significantly larger B-cell follicle.
For an appropriate acquired cellular immune response, it is important for T cells to survey secondary lymphoid organs thereby allowing contact with antigen-presenting dendritic cells that colocalize there.42-46 Such a nodal survey by T cells may be mediated by CCL21. In vitro evidence from us and others suggest that CCL21 is an extremely potent chemoattractant for normal human T cells, especially naïve T cells.33-36,61 In addition, data show that CCL21 is expressed in node endothelial cells and induces rapid adhesion of human T cells to endothelial ICAM-1.37-41 Lastly, the plt mouse mutation and the CCR7-deficient mouse model show disordered trafficking of T cells into nodal tissue.47-53 In this study transgenic mice were generated that placed the expression of mCCL21 under the control of the T cell-specific lck proximal promoter. Overexpression of CCL21 in T cells will abrogate any CCL21 chemokine concentration gradient that T cells might experience. Overexpression of mCCL21 by T cells in the lck/lck-mCCL21 transgenic mice resulted in defects in CCR7 ligand chemotaxis, the concentration of lymphocyte subpopulations in lymphoid structures, and nodal architecture. Deficiencies in transgenic CD3+/CD62L+ splenocyte and thymocyte chemotaxis in response to both mCCL21 and mCCL19 but not mCXCL12 were also observed. Because CCL21 and CCL19 share the CCR7 receptor, and CXCL12 interacts with CXCR4, it implies that this finding is receptor-specific. One possible mechanism for this chemotactic inhibition is that T-cell overexpression of CCL21 down-regulates the CCR7 receptor, thereby reducing its ability to respond to a CC21 gradient. This down-regulation could have occurred by either intracellular binding of the CCR7 receptor before it reached the surface or by endocytosis of the CCR7 receptor at the surface via clatharin- coated pits after it was bound by the secreted overexpressed CCL21. A second mechanism for this inhibition of cell migration is the destruction of a directional chemokine gradient to which the T cell would normally respond. Transgenic T cells would experience increased CCL21 protein levels completely surrounding the cell and no directional concentration gradient would exist. Adhesion mediated by CXCL12, CCL21, and CCL19 was unchanged in transgenic mice when compared to normal mice. Because T cells require CCL21 for the activation of integrins on their cell surface for adhesion, it is possible that transgenic overexpression of CCL21 provided that signal constitutively. This would produce transgenic T cells that are already primed for adherence, similar to the T cells exposed to exogenous CCL21. Adhesion may be unchanged by the overexpression of CCL21 in T cells because adhesion does not require a gradient, like chemotaxis, but simply the local presence of the chemokine for activation of CCR7. However, this finding does imply that the CCR7 receptor is reaching the cell surface in the lck/lck-mCCL21 T cells, and that the CCL21 ligand is not binding the CCR7 receptor intracellularly. Flow cytometry data revealed a small but statistically significant increase in the percent of CD62L+ T cells in the spleen and lymph nodes of lck/lck-mCCL21 transgenic mice when compared to WT/WT mice. One possible mechanism for this defect is that CD62L+ node-homing T cells are more sensitive to the CCL21 gradients. Thus, if any gradient in lymphoid tissue still existed, this subtype of T cell would sense it. Another possibility is that the mass of T cells collecting near the HEV produce large quantities of CCL21, resulting in a new relative gradient that circulating T cells preferentially sense, and localize there. This potential mechanism could also explain the abnormal node architecture observed in the transgenic mice. Examining nodal B- and T-cell architecture with immunohistology found that the transgenic mouse had a reduction in the size of the T-cell zone and an enlargement of the B-cell follicle. This is in contrast to the CCR7 knockout mice, where there was a lack of T-cell zones and B-cell follicles in the nodal architecture.55 Thus, modifying the CCL21 gradient a T cell would experience is not equivalent to deleting the CCR7 receptor. Either there still exists some small nodal CCL21 local gradient in the transgenic mice that allows T cells to enter the node, or other chemokines play a compensatory role in directing nodal T-cell migration. However, the adhesion and chemotaxis data together supply a more
satisfying explanation for the node architecture data. T cells do not
distribute normally throughout the node The data presented here provide additional evidence that CCL21 concentration gradients play an important role intranodal T-cell migration. However, these data also imply that other properties of CCR7 ligands besides a chemotactic concentration gradient (eg, adhesion) are important in naïve T-cell trafficking. In addition, this study implies that pharmacologically saturating the CCR7 receptor may decrease T-cell chemotaxis sufficiently to disrupt T cell-mediated autoimmune diseases. These transgenic mice provide a model to validate that hypothesis.
Submitted June 6, 2001; accepted August 8, 2001.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Robert A. Hromas, Departments of Biochemistry/Molecular Biology and Hematology/Oncology, Indiana University Cancer Center, Indiana University School of Medicine, 1044 W Walnut, Bldg R4-202, Indianapolis, IN 46202; e-mail: rhromas{at}iupui.edu.
1. Oppenheim JJ. Overview of chemokines. Adv Exp Med Biol. 1993;351:183-186[Medline] [Order article via Infotrieve]. 2. Schall TJ, Bacon KB. Chemokines, leukocyte trafficking, and inflammation. Curr Opin Immunol. 1994;6:865-873[CrossRef][Medline] [Order article via Infotrieve]. 3. Rossi D, Zlotnik A. The biology of chemokines and their receptors. Annu Rev Immunol. 2000;18:217-242[CrossRef][Medline] [Order article via Infotrieve]. 4. Kim CH, Broxmeyer HE. Chemokines: signal lamps for trafficking of T and B cells for development and effector function. J Leukoc Biol. 1999;65:6-15[Abstract]. 5. Locati M, Murphy PM. Chemokines and chemokine receptors: biology and clinical relevance in inflammation and AIDS. Annu Rev Med. 1999;50:425-440[CrossRef][Medline] [Order article via Infotrieve]. 6. Baggiolini M. Chemokines and leukocyte traffic. Nature. 1998;392:565-568[CrossRef][Medline] [Order article via Infotrieve]. 7. Feng L. Role of chemokines in inflammation and immunoregulation. Immunol Res. 2000;21:203-210[CrossRef][Medline] [Order article via Infotrieve].
8.
Rollins BJ.
Chemokines.
Blood.
1997;90:909-928
9.
Murphy PM, Baggiolini M, Charo IF, et al.
International union of pharmacology, XXII: nomenclature for chemokine receptors.
Pharmacol Rev.
2000;52:145-176 10. Sallusto F, Mackay CR, Lanzavecchia A. The role of chemokine receptors in primary, effector, and memory immune responses. Annu Rev Immunol. 2000;18:593-620[CrossRef][Medline] [Order article via Infotrieve]. 11. Rojo D, Suetomi K, Navarro J. Structural biology of chemokine receptors. Biol Res. 1999;32:263-272[Medline] [Order article via Infotrieve]. 12. Furie MB, Randolph GJ. Chemokines and tissue injury. Am J Pathol. 1995;146:1287-1301[Abstract]. 13. Baggiolini M, Dahinden CA. CC chemokines in allergic inflammation. Immunol Today. 1994;15:127-133[CrossRef][Medline] [Order article via Infotrieve]. 14. Strieter RM, Standiford TJ, Huffnagle GB, Colletti LM, Lukacs NW, Kunkel SL. "The good, the bad, and the ugly." The role of chemokines in models of human disease. J Immunol. 1996;156:3583-3586[Medline] [Order article via Infotrieve]. 15. Hosaka S, Akahoshi T, Wada C, Kondo H. Expression of the chemokine superfamily in rheumatoid arthritis. Clin Exp Immunol. 1994;97:451-457[Medline] [Order article via Infotrieve]. 16. Kasama T, Strieter RM, Lukacs NW, Lincoln PM, Burdick MD, Kunkel SL. Interleukin-10 expression and chemokine regulation during the evolution of murine type II collagen-induced arthritis. J Clin Invest. 1995;95:2868-2876. 17. Kukielka GL, Youker KA, Michael LH, et al. Role of early reperfusion in the induction of adhesion molecules and cytokines in previously ischemic myocardium. Mol Cell Biochem. 1995;147:5-12[CrossRef][Medline] [Order article via Infotrieve].
18.
Massey KD, Strieter RM, Kunkel SL, Danforth JM, Standiford TJ.
Cardiac myocytes release leukocyte-stimulating factors.
Am J Physiol.
1995;269:H980-H987 19. Feng Y, Broder CC, Kennedy PE, Berger EA. HIV-1 entry cofactor: functional cDNA cloning of a seven-transmembrane, G protein-coupled receptor. Science. 1996;272:872-877[Abstract]. 20. Dragic T, Litwin V, Allaway GP, et al. HIV-1 entry into CD4+ cells is mediated by the chemokine receptor CC-CKR-5. Nature. 1996;381:667-673[CrossRef][Medline] [Order article via Infotrieve]. 21. Alkhatib G, Combadiere C, Broder CC, et al. CC CKR5: a RANTES, MIP-1alpha, MIP-1beta receptor as a fusion cofactor for macrophage-tropic HIV-1. Science. 1996;272:1955-1958[Abstract]. 22. Muller A, Homey B, Soto H, et al. Involvement of chemokine receptors in breast cancer metastasis. Nature. 2001;410:50-56[CrossRef][Medline] [Order article via Infotrieve]. 23. Laning J, Kawasaki H, Tanaka E, Luo Y, Dorf ME. Inhibition of in vivo tumor growth by the beta chemokine, TCA3. J Immunol. 1994;153:4625-4635[Abstract]. 24. Manome Y, Wen PY, Hershowitz A. Monocyte chemoattractant protein-1 (MCP-1) gene transduction: an effective tumor vaccine strategy for non-intracranial tumors. Cancer Immunol Immunother. 1995;41:227-235[Medline] [Order article via Infotrieve]. 25. Dilloo D, Bacon K, Holden W, et al. Combined chemokine and cytokine gene transfer enhances antitumor immunity. Nat Med. 1996;2:1090-1095[CrossRef][Medline] [Order article via Infotrieve]. 26. Rossi DL, Vicari AP, Franz-Bacon K, McClanahan TK, Zlotnik A. Identification through bioinformatics of two new macrophage proinflammatory human chemokines: MIP-3alpha and MIP-3beta. J Immunol. 1997;158:1033-1036[Abstract].
27.
Yoshida R, Imai T, Hieshima K, et al.
Molecular cloning of a novel human CC chemokine EBI1-ligand chemokine that is a specific functional ligand for EBI1, CCR7.
J Biol Chem.
1997;272:13803-13809
28.
Braun SE, Chen K, Foster RG, et al.
The CC chemokine CK beta-11/MIP-3 beta/ELC/Exodus 3 mediates tumor rejection of murine breast cancer cells through NK cells.
J Immunol
2000;164:4025-4031
29.
Hromas R, Cripe L, Hangoc G, Cooper S, Broxmeyer HE.
The Exodus subfamily of CC chemokines inhibit the proliferation of chronic myelogenous leukemia progenitors.
Blood.
2000;95:1506-1508
30.
Sharma S, Stolina M, Luo J, et al.
Secondary lymphoid tissue chemokine mediates T cell-dependent antitumor responses in vivo.
J Immunol.
2000;164:4558-4563
31.
Vicari AP, Ait-Yahia S, Chemin K, Mueller A, Zlotnik A, Caux C.
Antitumor effects of the mouse chemokine 6Ckine/SLC through angiostatic and immunological mechanisms.
J Immunol.
2000;165:1992-2000 32. Nomura T, Hasegawa H. Chemokines and anti-cancer immunotherapy: anti-tumor effect of EBI1-ligand chemokine (ELC) and secondary lymphoid tissue chemokine (SLC). Anticancer Res. 2000;20:4073-4080[Medline] [Order article via Infotrieve]. 33. Hromas R, Kim CH, Klemsz M, et al. Isolation and characterization of Exodus-2, a novel C-C chemokine with a unique 37-amino acid carboxyl-terminal extension. J Immunol. 1997;159:2554-2558[Abstract]. 34. Hedrick JA, Zlotnik A. Identification and characterization of a novel beta chemokine containing six conserved cysteines. J Immunol. 1997;159:1589-1593[Abstract].
35.
Nagira M, Imai T, Hieshima K, et al.
Molecular cloning of a novel human CC chemokine secondary lymphoid-tissue chemokine that is a potent chemoattractant for lymphocytes and mapped to chromosome 9p13.
J Biol Chem.
1997;272:19518-19524 36. Tanabe S, Lu Z, Luo Y, et al. Identification of a new mouse beta-chemokine, thymus-derived chemotactic agent 4, with activity on T lymphocytes and mesangial cells. J Immunol. 1997;159:5671-5679[Abstract].
37.
Campbell JJ, Hedrick J, Zlotnik A, Siani MA, Thompson DA, Butcher EC.
Chemokines and the arrest of lymphocytes rolling under flow conditions.
Science.
1998;279:381-384
38.
Gunn MD, Tangemann K, Tam C, Cyster JG, Rosen SD, Williams LT.
A chemokine expressed in lymphoid high endothelial venules promotes the adhesion and chemotaxis of naive T lymphocytes.
Proc Natl Acad Sci U S A.
1998;95:258-263
39.
Tangemann K, Gunn MD, Giblin P, Rosen SD.
A high endothelial cell-derived chemokine induces rapid, efficient, and subset-selective arrest of rolling T lymphocytes on a reconstituted endothelial substrate.
J Immunol.
1998;161:6330-6337
40.
Campbell JJ, Bowman EP, Murphy K, et al.
6-C-kine (SLC), a lymphocyte adhesion-triggering chemokine expressed by high endothelium, is an agonist for the MIP-3beta receptor CCR7.
J Cell Biol.
1998;141:1053-1059
41.
Stein JV, Rot A, Luo Y, et al.
The CC chemokine thymus-derived chemotactic agent 4 (TCA-4, secondary lymphoid tissue chemokine, 6Ckine, Exodus-2) triggers lymphocyte function-associated antigen 1-mediated arrest of rolling T lymphocytes in peripheral lymph node high endothelial venules.
J Exp Med.
2000;191:61-76 42. Hedrick JA, Zlotnik A. Chemokines and chemokine receptors in T-cell development. Chem Immunol. 1999;72:57-68[Medline] [Order article via Infotrieve]. 43. Campbell JJ, Butcher EC. Chemokines in tissue-specific and microenvironment-specific lymphocyte homing. Curr Opin Immunol. 2000;12:336-341[CrossRef][Medline] [Order article via Infotrieve].
44.
Cyster JG.
Chemokines and cell migration in secondary lymphoid organs.
Science.
1999;286:2098-2102 45. Cyster JG. Leukocyte migration: scent of the T zone. Curr Biol. 2000;10:R30-R33[CrossRef][Medline] [Order article via Infotrieve].
46.
von Andrian UH, Mackay CR.
T-cell function and migration. Two sides of the same coin.
N Engl J Med.
2000;343:1020-1034 47. Nakano H, Tamura T, Yoshimoto T, et al. Genetic defect in T lymphocyte-specific homing into peripheral lymph nodes. Eur J Immunol. 1997;27:215-221[Medline] [Order article via Infotrieve].
48.
Nakano H, Mori S, Yonekawa H, Nariuchi H, Matsuzawa A, Kakiuchi T.
A novel mutant gene involved in T-lymphocyte-specific homing into peripheral lymphoid organs on mouse chromosome 4.
Blood.
1998;91:2886-2895
49.
Gunn MD, Kyuwa S, Tam C, et al.
Mice lacking expression of secondary lymphoid organ chemokine have defects in lymphocyte homing and dendritic cell localization.
J Exp Med.
1999;189:451-460
50.
Vassileva G, Soto H, Zlotnik A, et al.
The reduced expression of 6Ckine in the plt mouse results from the deletion of one of two 6Ckine genes.
J Exp Med.
1999;190:1183-1188
51.
Luther SA, Tang HL, Hyman PL, Farr AG, Cyster JG.
Coexpression of the chemokines ELC and SLC by T zone stromal cells and deletion of the ELC gene in the plt/plt mouse.
Proc Natl Acad Sci U S A.
2000;97:12694-12699
52.
Nakano H, Gunn MD.
Gene duplications at the chemokine locus on mouse chromosome 4: multiple strain-specific haplotypes and the deletion of secondary lymphoid-organ chemokine and EBI-1 ligand chemokine genes in the plt mutation.
J Immunol.
2001;166:361-369
53.
Mori S, Nakano H, Aritomi K, Wang CR, Gunn MD, Kakiuchi T.
Mice lacking expression of the chemokines CCL21-Ser and CCL19 (plt mice) demonstrate delayed but enhanced T cell immune responses.
J Exp Med.
2001;193:207-218 54. Forster R, Schubel A, Breitfeld D, et al. CCR7 coordinates the primary immune response by establishing functional microenvironments in secondary lymphoid organs. Cell. 1999;99:23-33[CrossRef][Medline] [Order article via Infotrieve]. 55. Chaffin KE, Beals CR, Wilkie TM, Forbush KA, Simon MI, Perlmutter RM. Dissection of thymocyte signaling pathways by in vivo expression of pertussis toxin ADP-ribosyltransferase. EMBO J 1990;9:3821-3829[Medline] [Order article via Infotrieve].
56.
Shimizu C, Kawamoto H, Yamashita M, et al.
Progression of T cell lineage restriction in the earliest subpopulation of murine adult thymus visualized by the expression of lck proximal promoter activity.
Int Immunol.
2001;13:105-117 57. Hogan B. Manipulating the Mouse Embryo. Plainview, NY: Cold Spring Harbor Laboratory Press; 1994:217-248. 58. Abbitt KB, Rainger GE, Nash GB. Effects of fluorescent dyes on selectin and integrin-mediated stages of adhesion and migration of flowing leukocytes. J Immunol Methods. 2000;239:109-119[CrossRef][Medline] [Order article via Infotrieve]. 59. Frevert CW, Wong VA, Goodman RB, Goodwin R, Martin TR. Rapid fluorescence-based measurement of neutrophil migration in vitro. J Immunol Methods. 1998;213:41-52[CrossRef][Medline] [Order article via Infotrieve]. 60. Gildea JJ, Harding MA, Gulding KM, Theodorescu D. Transmembrane motility assay of transiently transfected cells by fluorescent cell counting and luciferase measurement. Biotechniques. 2000;29:81-86[Medline] [Order article via Infotrieve]. 61. Christopherson K 2nd, Brahmi Z, Hromas R. Regulation of naive fetal T-cell migration by the chemokines Exodus-2 and Exodus-3. Immunol Lett. 1999;69:269-273[CrossRef][Medline] [Order article via Infotrieve].
62.
Hromas R, Orazi O, Neiman R, et al.
Hematopoietic lineage and stage-restricted expression of the ETS oncogene family member PU.1.
Blood.
1993;82:2998-3004
© 2001 by The American Society of Hematology.
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 H. Unsoeld, K. Mueller, U. Schleicher, C. Bogdan, J. Zwirner, D. Voehringer, and H. Pircher Abrogation of CCL21 chemokine function by transgenic over-expression impairs T cell immunity to local infections Int. Immunol., November 1, 2007; 19(11): 1281 - 1289. [Abstract] [Full Text] [PDF]
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 S. Basu and H. E. Broxmeyer Transforming growth factor-{beta}1 modulates responses of CD34+ cord blood cells to stromal cell-derived factor-1/CXCL12 Blood, July 15, 2005; 106(2): 485 - 493. [Abstract] [Full Text] [PDF]
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 Y. Guo, R. Costa, H. Ramsey, T. Starnes, G. Vance, K. Robertson, M. Kelley, R. Reinbold, H. Scholer, and R. Hromas The embryonic stem cell transcription factors Oct-4 and FoxD3 interact to regulate endodermal-specific promoter expression PNAS, March 19, 2002; 99(6): 3663 - 3667. [Abstract] [Full Text] [PDF]
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Top
Abstract
Introduction
-actin antibody was used to confirm equal loading.
(C) shows a statistically
significant increase in CD62L+ T cells in the spleen,
thymus, and lymph nodes of lck/lck-mCCL21 transgenic mice, when
compared to normal WT mice. Corresponding decreases in
CD62L
they exit the HEV normally but
then fail to migrate properly throughout the node. Yet flow cytometry
data indicated that the number of T cells relative to B cells remains
unchanged in the lck/lck-CCL21 transgenic mice compared to normal mice.
A potential explanation for this is that T-cell/endothelial cell
adhesion (normal in these transgenic mice) is the key requirement for
HEV node entry,