Biomedical subjects

D D Taub

Publications and source records attributed to D D Taub.

At least 37 records · Page 2Linked to original sources

PubMed1999

Cocaine enhances brain endothelial adhesion molecules and leukocyte migration.

Leukocyte infiltration of cerebral vessels in cocaine-associated vasculopathy suggests that cocaine may enhance leukocyte migration. We have investigated cocaine's effects on leukocyte adhesion in human brain microvascular endothelial cell (BMVEC) cultures and monocyte migration in an in vitro blood-brain barrier (BBB) model constructed with BMVEC and astrocytes. Cocaine (10(-5) to 10(-9) M) enhanced adhesion of monocytes and neutrophils to BMVEC. In the BBB model, cocaine (10(-4) to 10(-8) M) enhanced monocyte transmigration. Cocaine increased expression of endothelial adhesion molecules, intercellular adhesion molecule-1 (ICAM-1, CD54), vascular cell adhesion molecule-1 (VCAM-1), and endothelial leukocyte adhesion molecule-1 (ELAM-1) on BMVEC. The peak effect on ICAM-1 expression was between 6 and 18 h after treatment. ICAM-1 was increased by cocaine in BMVEC, but not in human umbilical vein endothelial cells, and the enhancement was greater in a coculture of BMVEC with monocytes. ICAM-1 expression was enhanced by a transcriptional mechanism. Polymyxin B inhibited up-regulation of adhesion molecules by LPS but not by cocaine. In LPS-activated BMVEC/monocyte coculture, cocaine increased secretion of tumor necrosis factor-alpha and interleukin-6. Taken together, these findings indicate that cocaine enhances leukocyte migration across the cerebral vessel wall, in particular under inflammatory conditions, but the effects are variable in different individuals. Cocaine's effects are exerted through a cascade of augmented expression of inflammatory cytokines and endothelial adhesion molecules. These could underlie the cerebrovascular complications of cocaine abuse.

Astrocytes↗

PubMed1999

Treatment with tumor necrosis factor-alpha and granulocyte-macrophage colony-stimulating factor increases epidermal Langerhans' cell numbers in cancer patients.

Dendritic cells (DCs) initiate primary and stimulate secondary T-cell responses. We conducted a phase I trial of tumor necrosis factor (TNF-alpha) and granulocyte-macrophage colony-stimulating factor (GM-CSF) in patients with cancer to increase DCs in peripheral blood or skin based on in vitro data that showed that CD34(+) hematopoietic precursors require these cytokines to mature into functional antigen-presenting DCs. Eleven patients were treated for 7 days with GM-CSF, 125 microg/m(2) twice daily as subcutaneous injections, and TNF-alpha as a continuous infusion at dose levels of 25, 50, or 100 microg/m(2)/day. The maximum tolerated dose of TNF-alpha was 50 microg/m(2)/day with this dose of GM-CSF; dose-limiting toxicities occurred in both patients treated with 100 microg/m(2)/day. One became thrombocytopenic and the other had transient confusion. Epidermal Langerhans' cells were quantitated by S100 staining of skin biopsies and DC precursors in peripheral blood by colony-forming unit dendritic (CFU-dendritic) assays. S100-positive cells in the epidermis doubled after treatment (2.55 S100(+) cells/high-power field before treatment to 6.05 after treatment, p = 0.029). CFU-dendritic in peripheral blood increased after treatment in 3 colorectal cancer patients but not in 3 patients with melanoma. CD11c(+) or CD123(+), HLA-DR(bright), lineage-negative dendritic cell precursors were not increased in peripheral blood mononuclear cells. This trial demonstrates that treatment with TNF-alpha and GM-CSF can increase the number of DCs in the skin and the number of dendritic cell precursors in the blood of some patients with cancer. This approach may increase the efficacy of vaccination to tumor antigens in cancer patients.

Adult↗

PubMed1998

Differential effects of the absence of interferon-gamma and IL-4 in acute graft-versus-host disease after allogeneic bone marrow transplantation in mice.

Graft-versus-host disease (GVHD), in which immunocompetent donor cells attack the host, remains a major cause of morbidity after allogeneic bone marrow transplantation (BMT). To understand the role of cytokines in the pathobiology of GVHD, we used cytokine knockout (KO) mice as a source of donor T cells. Two different MHC-disparate strain combinations were examined: BALB/c (H2(d)) donors into lethally irradiated C57BL/6 (H2(b)) recipients or C57BL/6 (H2(b)) donors into B10.BR (H2(k)) recipients. Donor cells were from mice in which either the interferon-gamma (IFN-gamma) or the IL-4 gene was selectively disrupted to understand the role of these cytokines in acute GVHD. In both strain combinations the same pattern was noted with regard to GVHD onset and morbidity. All mice exhibited the classic signs of acute GVHD: weight loss with skin, gut, and liver pathology resulting in morbidity and mortality. Surprisingly, donor cells obtained from mice lacking IFN-gamma gave rise to accelerated morbidity from GVHD when compared with cells from wild-type control donors. Similar results were obtained using normal donors when neutralizing antibodies to IFN-gamma were administered immediately after the BMT. These results suggest that IFN-gamma plays a role in protection from acute GVHD. In marked contrast, cells obtained from IL-4 KO mice resulted in protection from GVHD compared with control donors. Splenocytes from IFN KO mice stimulated with a mitogen proliferated to a significantly greater extent and produced more IL-2 compared with splenocytes obtained from IL-4 KO or control mice. Additionally, there was increased IL-2 production in the spleens of mice undergoing GVHD using IFN-gamma KO donors. These results therefore indicate, with regard to the TH1/ TH2 cytokine paradigm, the absence of a TH1-type cytokine can be deleterious in acute GVHD, whereas absence of a TH2 cytokine can be protective.

Animals↗

PubMed1998

Molecular cloning of FKHRL1P2, a member of the developmentally regulated fork head domain transcription factor family.

Here we report the expression of a fork head domain protein in human T helper cells. We cloned and characterized a fork head cDNA from human T helper cell mRNA using differential display RT-PCR. The cDNA contains a 546-nucleotide (nt) open reading frame (ORF) that codes for the carboxyl-terminal 180 amino acids (aa) of the recently identified fkhrl1 gene. This ORF does not contain the characteristic DNA-binding domain found in members of the forkhead protein family. In-vitro transcription/translation of this cDNA expressed a protein of approximately 20 kDa. We have generated antibodies that specifically immunoprecipitated the in-vitro-translated 20-kDa protein. This antibody also recognizes in human T lymphocytes a 70-kDa protein corresponding in size to that predicted for the fkhrl1 gene product. The mRNA levels for fkhrl1 is elevated in T helper-induced lymphocytes in comparison to PHA-stimulated T lymphocytes. Further characterization of FKHRL1 and its related family members should shed light on the transcriptional mechanisms of this fork head gene subfamily and their role in T helper cell differentiation and regulation of cell growth.

Amino Acid Sequence↗

PubMed1998

Opiates transdeactivate chemokine receptors: delta and mu opiate receptor-mediated heterologous desensitization.

An intact chemotactic response is vital for leukocyte trafficking and host defense. Opiates are known to exert a number of immunomodulating effects in vitro and in vivo, and we sought to determine whether they were capable of inhibiting chemokine-induced directional migration of human leukocytes, and if so, to ascertain the mechanism involved. The endogenous opioid met-enkephalin induced monocyte chemotaxis in a pertussis toxin-sensitive manner. Met-enkephalin, as well as morphine, inhibited IL-8-induced chemotaxis of human neutrophils and macrophage inflammatory protein (MIP)-1alpha, regulated upon activation, normal T expressed and secreted (RANTES), and monocyte chemoattractant protein 1, but not MIP-1beta-induced chemotaxis of human monocytes. This inhibition of chemotaxis was mediated by delta and micro but not kappa G protein-coupled opiate receptors. Calcium flux induced by chemokines was unaffected by met-enkephalin pretreatment. Unlike other opiate-induced changes in leukocyte function, the inhibition of chemotaxis was not mediated by nitric oxide. Opiates induced phosphorylation of the chemokine receptors CXCR1 and CXCR2, but neither induced internalization of chemokine receptors nor perturbed chemokine binding. Thus, inhibition of chemokine-induced chemotaxis by opiates is due to heterologous desensitization through phosphorylation of chemokine receptors. This may contribute to the defects in host defense seen with opiate abuse and has important implications for immunomodulation induced by several endogenous neuropeptides which act through G protein-coupled receptors.

Cells, Cultured↗

PubMed1998

Identification and characterization of small molecule functional antagonists of the CCR1 chemokine receptor.

The CC chemokines macrophage inflammatory protein-1alpha (MIP-1alpha) and RANTES (regulated on activation normal T cell expressed) have been implicated in rheumatoid arthritis and multiple sclerosis. Since their effects are mediated through the CCR1 chemokine receptor, we set up a small molecule CCR1 antagonist program to search for inhibitors. Through high capacity screening we discovered a number of 4-hydroxypiperidine compounds with CCR1 antagonist activity and report their synthesis and in vitro pharmacology here. Scatchard analysis of the competition binding data revealed that the compounds had Ki values ranging from 40 to 4000 nM. The pharmacological profile of the most potent member of this series, compound 1 (2-2-diphenyl-5-(4-chlorophenyl)piperidin-lyl)valeronitri te), was further evaluated. Compound 1 showed concentration-dependent inhibition of MIP-1alpha-induced extracellular acidification and Ca2+ mobilization demonstrating functional antagonism. When given alone, the compound did not elicit any responses, indicating the absence of intrinsic agonist activity. Compound 1 inhibited MIP-1alpha- and RANTES-induced migration in peripheral blood mononuclear cells in a dose-responsive manner. Selectivity testing against a panel of seven transmembrane domain receptors indicated that compound 1 is inactive on a number of receptors at concentrations up to 10 microM. This is the first description of CCR1 receptor antagonists that may be useful in the treatment of chronic inflammatory diseases involving MIP-1alpha, RANTES, and CCR1.

Arthritis, Rheumatoid↗

PubMed1998

Novel role of transmembrane SCF for mast cell activation and eotaxin production in mast cell-fibroblast interactions.

Mast cell activation can be induced by multiple mechanisms, including IgE-, complement-, and stem cell factor (SCF)-mediated pathways. In addition, the interaction of mast cells with particular cell populations, such as fibroblasts, have also demonstrated increased mast cell reactivity. In these studies, we have investigated the role of fibroblast-mast cell interaction for induction of histamine release and chemokine production and the specific role of SCF during this interaction. Primary pulmonary fibroblast cell lines were grown in culture and used throughout these studies. Mast cells were grown in parallel with fibroblasts by incubation of bone marrow cells with SCF and IL-3. During mast cell-fibroblast coculture, increased histamine release could be attenuated either by separation of the cell populations using a Trans-Well setup, which did not allow cellular contact, or by specific anti-SCF Ab. In addition, a significant increase in eotaxin, a potent eosinophil-specific C-C chemokine, was also observed during fibroblast-mast cell interaction. The production of eotaxin was cell contact dependent and could be inhibited using an anti-SCF Ab or specific antisense therapy. SCF was constitutively produced from fibroblasts in its transmembrane form and could be induced by TNF. SCF-coated plates induced significant mast cell-derived eotaxin production, whereas soluble SCF induced little or no eotaxin, suggesting a necessity for receptor cross-linking for activation. These studies indicate that fibroblast-mast cell contact plays a role in exacerbation of histamine release and eotaxin production.

Animals↗

PubMed1998

Opiate inhibition of chemokine-induced chemotaxis.

Chemokines consist of a family of 8-16-kDa cytokines that are generated very early in a wide variety of inflammatory responses and attract leukocytes to local sites. At nanomolar concentrations chemokines initiate signal transduction and activate leukocytes through seven transmembrane receptors (STM), but higher micromolar doses result in homologous desensitizing effects. On the basis of reports that opiates have anti-inflammatory effects and also use STM, we have investigated the possibility that they may cross-desensitize the response of leukocytes to chemokines. We have confirmed previous observations that met-enkephalin (MET) is chemotactic for human peripheral blood monocytes. Furthermore, we observed that preincubation of monocytes or neutrophils with MET or morphine prevented their subsequent chemotaxis in response to chemokines (MIP1 alpha or IL-8). However, MET did not inhibit the chemotactic response of PMN to NAP-2, a homologous chemokine that is less potent than IL-8 but cannot be desensitized. The inhibitory effect of opiates on chemokine-induced chemotaxis was antagonized by naloxone. Since MIP-1 alpha and IL-8, unlike NAP-2, have the capacity to desensitize leukocytes, it is possible that opiates, by desensitizing some chemokine responses, can suppress inflammatory reactions.

Animals↗

PubMed1998

Suppression of graft-versus-host disease and amplification of graft-versus-tumor effects by activated natural killer cells after allogeneic bone marrow transplantation.

Bone marrow transplantation (BMT) is currently used for the treatment of a variety of neoplastic diseases. However, significant obstacles limiting the efficacy of allogeneic BMT are the occurrence of graft-versus-host disease (GvHD) and tumor relapse. Natural killer (NK) cells exert a variety of immunologic and homoeostatic functions. We examined whether adoptive transfer of activated NK cells of donor type would prevent GvHD after allogeneic BMT in mice. Lethally irradiated C57BL/6 (H-2(b)) mice, were transplanted with MHC incompatible BALB/c (H-2(d)) bone marrow cells and spleen cells and rapidly succumbed to acute GvHD. In contrast, mice that also received activated NK cells of donor type exhibited significant increases in survival. In determining the mechanism by which the NK cells prevented GvHD, mice were concurrently treated with a neutralizing antibodies to the immunosuppressive cytokine TGFbeta. Anti-TGFbeta completely abrogated the protective effects of the activated donor NK cells indicating that TGFbeta plays an important role in the prevention of GvHD by NK cells. We then examined whether activated NK cells of donor type after allogeneic BMT would induce graft-versus-tumor (GvT) effects without GvHD in mice bearing a murine colon adenocarcinoma (MCA-38). 10 d after receiving the tumor, in which the mice had demonstrable lung metastases, recipients received an allogeneic BMT with or without activated NK cells. Administration of activated NK cells resulted in significant GvT effects after allogeneic BMT as evidenced by increases in median survival and fewer lung metastasis. No evidence of GVHD was detected compared with recipients receiving spleen cells alone which also developed fewer lung metastases but in which all had succumbed to GVHD. Thus, our findings suggest that adoptive immunotherapy using activated donor NK cells combined with allogeneic BMT inhibits GvHD and promotes GvT in advanced tumor-bearing mice. These results also suggest that GvT and GvHD can be dissociable phenomena.

Adenocarcinoma↗

PubMed1998

CD8+ myelin peptide-specific T cells can chemoattract CD4+ myelin peptide-specific T cells: importance of IFN-inducible protein 10.

The demyelination process that occurs in the central nervous system (CNS) of patients with multiple sclerosis (MS) is due, in part, to an inflammatory response in which CD4+ and CD8+ T cells and macrophages infiltrate white matter. While it is thought that the inflammatory and demyelination process in MS is the product of Th1-associated cytokines secreted by CD4+ myelin protein-specific T cells present in the CNS, the mechanisms that are responsible for the recruitment and maintenance of these myelin-reactive CD4+ T cells in the CNS have not been elucidated. We have shown previously that CD8+ CTL that recognize peptides derived from sequences of the myelin proteolipid protein (PLP) presented by HLA class I molecules can be generated in vitro, and that these PLP-specific CD8+ CTL secrete the proinflammatory chemokines macrophage-inflammatory protein-1alpha and -1beta, IL-16, and IP-10. In this study, we demonstrate that soluble products of these PLP-specific CD8+ CTL can chemoattract CD4+ T cells that are specific for a myelin basic protein peptide and a PLP peptide, and that the majority of this chemotactic activity is mediated by IFN-inducible protein 10. These results demonstrate that PLP-specific CD8+ T cells can play a role in the recruitment and retention of myelin-derived peptide-specific CD4+ T cells, and indicate that they may play a proinflammatory role in the pathogenesis of MS.

Amino Acid Sequence↗

PubMed1998

Sticky and smelly issues: lessons on tumour cell and leucocyte trafficking, gene and immunotherapy of cancer.

The Second Meeting of the British Society for Immunology Tumour Immunology Affinity Group (TIAG) took place at King's College (London, UK) on 17-18 June 1997 and brought together over 100 tumour immunologists from the UK and abroad. In contrast to previous meetings the focus of the meeting was on the role of adhesion in immunosurveillance and tumour dissemination. In addition, recent achievements in the areas of chemokines, cytotoxic T-lymphocyte (CTL) and natural killer (NK) cells, co-stimulation, gene and adoptive immunotherapy were also addressed. The purpose of this report is to outline current trends in tumour immunology.

Animals↗

PubMed1998

Similar levels of human immunodeficiency virus type 1 replication in human TH1 and TH2 clones.

Studies on the development and function of CD4+ TH1 and TH2 cells during the progression to AIDS may increase the understanding of AIDS pathogenesis. The preferential replication of human immunodeficiency virus (HIV) in either TH1 or TH2 cells could alter the delicate balance of the immune response. TH1 (gamma interferon [IFN-gamma] positive, interleukin-4 [IL-4] and IL-5 negative) and TH2 (IFN-gamma negative, IL-4 and IL-5 positive) clones, developed from several healthy donors, pedigreed by reverse transcriptase PCR (RT-PCR) and enzyme linked immunosorbent assay have similar levels of cell surface expression of CD4 and several chemokine receptor cofactors necessary for viral entry. After activation by specific antigens and infection with T-cell-tropic strains of HIV type 1 (HIV-1), TH1 and TH2 clones showed similar levels of viral entry and reverse transcription. At days 3 through 14 postinfection, HIV replicated to similar levels in several TH1 and TH2 clones as measured by release of HIV p24 and total number of copies of gag RNA/total cell RNA as measured by RT-PCR. When values were normalized for viable cell number in three clones of each type, there was up to twofold more HIV RNA in TH1 than TH2 cells. In addition, several primary monocytotropic HIV-1 strains were able to replicate to similar levels in TH1 and TH2 cells. These studies suggest that the importance of TH1 and TH2 subsets in AIDS pathogenesis transcends clonal differences in their ability to support HIV replication.

CD4 Antigens↗

PubMed1998

Infection with human immunodeficiency virus type 1 upregulates DNA methyltransferase, resulting in de novo methylation of the gamma interferon (IFN-gamma) promoter and subsequent downregulation of IFN-gamma production.

The immune response to pathogens is regulated by a delicate balance of cytokines. The dysregulation of cytokine gene expression, including interleukin-12, tumor necrosis factor alpha, and gamma interferon (IFN-gamma), following human retrovirus infection is well documented. One process by which such gene expression may be modulated is altered DNA methylation. In subsets of T-helper cells, the expression of IFN-gamma, a cytokine important to the immune response to viral infection, is regulated in part by DNA methylation such that mRNA expression inversely correlates with the methylation status of the promoter. Of the many possible genes whose methylation status could be affected by viral infection, we examined the IFN-gamma gene as a candidate. We show here that acute infection of cells with human immunodeficiency virus type 1 (HIV-1) results in (i) increased DNA methyltransferase expression and activity, (ii) an overall increase in methylation of DNA in infected cells, and (iii) the de novo methylation of a CpG dinucleotide in the IFN-gamma gene promoter, resulting in the subsequent downregulation of expression of this cytokine. The introduction of an antisense methyltransferase construct into lymphoid cells resulted in markedly decreased methyltransferase expression, hypomethylation throughout the IFN-gamma gene, and increased IFN-gamma production, demonstrating a direct link between methyltransferase and IFN-gamma gene expression. The ability of increased DNA methyltransferase activity to downregulate the expression of genes like the IFN-gamma gene may be one of the mechanisms for dysfunction of T cells in HIV-1-infected individuals.

CD4-Positive T-Lymphocytes↗

PubMed1998

Induction of melanoma antigen-specific cytotoxic T lymphocytes in vitro by stimulation with B7-expressing human melanoma cell lines.

Crosslinking of CD28 receptors on resting T lymphocytes by B7 costimulatory molecules expressed by antigen-presenting cells (APCs) plays a critical role in T-cell activation. Human melanomas express major histocompatibility complex (MHC)-restricted tumor-associated antigens that can be recognized by cytotoxic T lymphocytes (CTL), yet they remain poorly immunogenic. One mechanism for the failure of T-cell response is the lack of expression of costimulatory molecules by human melanoma cells. We have transfected the B7-1 gene into three HLA-A2-expressing human melanoma cell lines, and studied their capacity to stimulate primary human T cells. B7-expressing melanoma cells were excellent inducers of T-cell proliferation, cytokine production, and cytolytic activity in allogeneic mixed lymphocyte cultures through a process dependent on the function of the T-cell receptor as well as interactions between B7:CD28, CD2:LFA-3, and LFA-1:ICAM-1. Subset analysis demonstrated that CD4+ T cells or addition of exogenous interleukin-2 was required for the induction of CD8+ CTL. Untransfected parental melanoma cells were inert as APCs in these cultures. Rotating stimulation of T cells with the three B7-expressing cell lines led to the generation of T-cell lines that were cytolytic for HLA-A2+ melanoma cells and other HLA-A2+ targets that were pulsed with HLA-A2-restricted MART-1 peptides. These data demonstrate that expression of B7-1 by human melanoma cells converts them into effective APCs for the in vitro induction of MHC-restricted, melanoma-specific CTL.

Antigen-Presenting Cells↗

PubMed1997

Human T cell leukemia virus type I (HTLV-I)-specific CD8+ CTL clones from patients with HTLV-I-associated neurologic disease secrete proinflammatory cytokines, chemokines, and matrix metalloproteinase.

Human T cell leukemia virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is a chronic, progressive neurologic disease characterized by marked degeneration of the spinal cord and the presence of infiltrating CD8+ T cells and macrophages. HAM/TSP patients have very high frequencies of HTLV-I-specific CD8+ CTL in peripheral blood and in cerebrospinal fluid. In this study, we show that HAM/TSP patients also have elevated levels of peripheral blood CD8+ T cells that produce intracellular IFN-gamma. To address the potential role of soluble mediators secreted by CD8+ T cells in the pathogenesis of HAM/TSP, we have analyzed the capacity of a panel of nine HTLV-I-specific CD8+ CTL clones derived from three HAM/TSP patients to secrete cytokines, chemokines, and matrix metalloproteinases. The results demonstrate that the majority of these CTL clones secrete IFN-gamma, TNF-alpha, macrophage-inflammatory protein-1alpha and -1beta, IL-16, and matrix metalloproteinase-9. These findings indicate that HTLV-I-specific CD8+ CTL are an important source of proinflammatory soluble mediators that may contribute significantly to the pathogenesis of HAM/TSP.

Chemokines↗

PubMed1997

Molecular cloning and characterization of a cDNA, CHEMR1, encoding a chemokine receptor with a homology to the human C-C chemokine receptor, CCR-4.

Chemokines refer to a rapidly expanding family of small cytokines whose primary function is recruitment of leukocytes to inflammatory sites. These are known to bind to seven-transmembrane-domain containing receptors. A cDNA clone, CHEMR1, resembling the typical G protein-coupled receptor, was isolated from a mouse cytotoxic T-lymphocyte (CTL) library. Northern blot analysis in mouse cell lines suggests that its expression is found in a variety of cells, including T cells, B cells, and macrophages. The CHEMR1 gene Scya3r2 is a single-copy gene whose open reading frame may be in a single exon and maps to the distal region of mouse Chr 9 where the mouse macrophage inflammatory protein-1alpha (MIP-1alpha) receptor gene Scya3r and two related C-C chemokine receptor-like genes reside. Amino acid sequence comparison shows that CHEMR1 is 84% identical to human CCR-4, indicating that CHEMR1 is likely to be a mouse CCR-4. Binding assays using 125I-labeled C-C chemokines in mammalian cells indicated that CHEMR1 did not bind MIP-1alpha, RANTES, or MIP-1beta, whereas CCR-1 binds MIP-1alpha and RANTES. Our result is different from the reported properties of human CCR-4. This suggests that CHEMR1 may be a receptor for unidentified C-C chemokine or a low-affinity receptor for MIP-1alpha.

Amino Acid Sequence↗

PubMed1997

Chemokines induce migrational responses in human breast carcinoma cell lines.

Chemokines have been shown to chemoattract and activate different leukocyte populations. Here we report the in vitro effect of macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, regulated on activation, normal T-cells, expressed and secreted (RANTES), monocyte chemotactic protein-1 (MCP-1), interleukin-8 (IL-8), interferon inducible protein-10 (IP-10), neutrophil-activating peptide-2 (NAP-2), growth-related protein (GRO)-alpha and GRO-gamma, on the migration of 3 human breast carcinoma cell lines, MCF-7, T47D and ZR-75-1, using a microchemotaxis chamber to assess migration across fibronectin-coated polycarbonate membranes. MCF-7 cells responded chemotactically to all chemokines tested in a pattern which was dose and time dependent, although responses to the different chemokines were variable. ZR-75-1 responded to MIP-1beta and GRO-alpha, giving maximum migration indices of 3.7 and 5.3, respectively, and exhibited a migratory response to MIP-1alpha, IL-8 and MCP-1 although to a lower degree. T47D was unresponsive to the chemokines tested, but both MCF-7 and T47D cells bound radiolabelled ligands with binding constants (Kd) ranging from 0.6 to 2.2 nM and 0.6 to 2.1 nM, respectively. The specificity of the chemotactic response of MCF-7 to MIP-1alpha and MIP-1beta was confirmed using chemokine-specific neutralising antibodies and heat denaturation, and was demonstrated to involve G protein and cyclic AMP signalling pathways. MIP-1beta and MIP-1alpha were shown to induce changes in the organisation of the actin cytoskeleton and the level of F-actin in MCF-7 cells, as determined using flow cytometric analysis and confocal microscopy. Our results show that breast carcinoma cells can respond to chemokines, and suggests a potential role for these molecules in the process of tumour cell migration, invasion and metastasis.

Actins↗

PubMed1997

Chemokine and matrix metalloproteinase secretion by myelin proteolipid protein-specific CD8+ T cells: potential roles in inflammation.

The demyelination process that occurs in the central nervous system of patients with multiple sclerosis (MS) is, in part, due to an inflammatory response in which CD4+ and CD8+ T cells and macrophages infiltrate white matter. Although many studies have characterized myelin protein-specific CD4+ T cells, we have demonstrated that CD8+ CTL specific for myelin peptides can be identified. In the present study, the potential roles of these CD8+ CTL in the generation of the inflammatory responses associated with MS have been investigated by measuring the capacity of these T cells to secrete the following proinflammatory chemokines: macrophage inflammatory protein-1alpha (MIP-1alpha) and MIP-1beta, lymphocyte chemoattractant factor (IL-16), IFN-inducible protein-10, as well as the proinflammatory enzymes of the matrix metalloproteinase family. The CD8+ CTL lines tested are derived from MS patients and are specific for two different myelin proteolipid protein-derived peptides presented by HLA-A2 and HLA-A3. All of the 17 CD8+ CTL lines secreted detectable amounts of MIP-1alpha and MIP-1beta. Nine of twelve CTL lines tested secreted IL-16, 10 of 12 lines tested secreted IFN-inducible protein-10, and 14 of 16 lines tested secreted matrix metalloproteinase-9. Collectively, these results indicate that myelin proteolipid protein peptide-specific CD8+ CTL may be an important source of proinflammatory soluble mediators that could promote and mediate the inflammatory response in MS demyelination.

Cell Line↗