Changes in chemokines and their receptors in blood during treatment with the TNF inhibitor infliximab in patients with rheumatoid arthritis.

OBJECTIVES: Chemokines are involved in leucocyte recruitment into inflammatory sites. The release of certain chemokines is augmented by tumour necrosis factor (TNF). Infliximab, a monoclonal antibody that blocks the effects of TNF, is used for treatment of rheumatoid arthritis (RA). The effect of TNF blockage on chemokines is not fully understood. The aim of this study was to analyse the effects on chemokines and their receptors on peripheral mononuclear cells of anti-TNF treatment in RA patients. METHOD: Twelve patients with established RA who started treatment with infliximab and nine patients with early RA treated with other anti-rheumatic drugs were followed clinically for 30 weeks and chemokine levels in blood samples were analysed along with chemokine receptor expression on the surface of T cells and monocytes. Nine healthy subjects were included as a control group. RESULTS: The chemokine CXCL10/IP-10 was significantly higher in RA patients than in healthy controls (p = 0.012). Two weeks after infliximab infusion, CXCL10/IP-10, CCL2/MCP-1, and CCL4/MIP-1ß had decreased significantly (p = 0.005, 0.037, and 0.028, respectively), and after 30 weeks of treatment, soluble CD26 was significantly increased (p = 0.050). Several chemokine receptors on T cells were elevated in RA patients at inclusion. The expression of CCR2 and CXCR1 on T cells decreased significantly after infliximab treatment. CONCLUSIONS: The chemokines CXCL10/IP-10, CCL2/MCP-1, and CCL4/MIP-1ß, mainly targeting the T-helper (Th)1 immune response, decreased after treatment with anti-TNF, suggesting a more pronounced effect on Th1 activity than on Th2-mediated response. Several chemokine receptors on blood T cells were elevated in RA patients, suggesting that they may be involved in the recruitment of T lymphocytes from the blood to affected tissues.

Receptor communication within the lymphocyte plasma membrane: a role for the thrombospondin family of matricellular proteins.

Lymphocytes, the principal cells of the immune system, carry out immune surveillance throughout the body by their unique capacity to constantly reposition themselves between a free-floating vascular state and a tissue state characterized by migration and frequent adhesive interactions with endothelial cells and components of the extracellular matrix. Therefore, mechanisms co-ordinating adhesion and migration with signals delivered through antigen recognition probably play a pivotal role for the regulation of lymphocyte behaviour and function. Endogenous thrombospondin-1 (TSP-1) seems to be the hub in such a mechanism for autocrine regulation of T cell adhesion and migration. TSP-1 functions as a mediator of cis interaction of vital receptors within the T lymphocyte plasma membrane, including integrins, low density lipoprotein receptor-related protein, calreticulin and integrin-associated protein.

Autoantibody formation in patients with rheumatoid arthritis treated with anti-TNF alpha.

BACKGROUND: Research on autoantibody formation in patients treated with TNF alpha inhibitors has produced contradictory results. OBJECTIVE: To study the prevalence of autoantibodies in patients with rheumatoid arthritis treated with the TNF alpha inhibitor infliximab. METHODS: 53 patients (48 female, 11 male) treated with infliximab for rheumatoid arthritis were followed for autoantibody production before treatment and after 14, 30, and 54 weeks. Six patients treated with etanercept were studied for comparison. The analyses included antibodies against nuclear antigens (ANA), extractable nuclear antigens, double stranded (ds)DNA (by ELISA, IIF on Crithidia luciliae for IgM and IgG, and Farr assay), nucleosomes, cardiolipin, smooth muscle, mitochondria, proteinase 3, and myeloperoxidase antigens. RESULTS: The number of patients treated with infliximab who developed antibodies against dsDNA of both IgG and IgM class (tested by IIF) increased significantly. The prevalence of patients positive for IgG class increased to 66% at 30 weeks and 45% at 54 weeks, and of IgM class to 85% and 70%, respectively. The titre and number of patients expressing antibodies against nucleosomes and ANA also increased significantly. The number of rheumatoid factor or anticardiolipin positive patients was stable and there was no increase in antibodies against the other antigens. A lupus-like syndrome was seen in one patient. No patient treated with etanercept developed any of these autoantibodies. CONCLUSIONS: Patients treated with infliximab may develop anti-dsDNA antibodies of both IgM and IgG class, anti-nucleosome antibodies, and ANA, with a gradual increase until 30 weeks.

Somatostatin is a specific inhibitor of SDF-1alpha-induced T cell infiltration.

The chemokine stromal cell-derived factor 1alpha (SDF-1alpha) is a potent stimulator of T cell infiltration into three-dimensional type I collagen matrices as demonstrated using T cells freshly isolated from blood and an activated T cell clone. The neuropeptide somatostatin selectively inhibits SDF-1alpha induced T cell infiltration by the same T cells including CD4 as well as CD8 positive cells, while somatostatin does not inhibit 'spontaneous' T cell infiltration. A number of other neuropeptides and opioids do not inhibit SDF-1alpha-induced T cell infiltration, indicating that the inhibitory effect is somatostatin-specific. The neuropeptide antagonist cyclosomatostatin abrogated the inhibitory effect of somatostatin on T cell infiltration, indicating that the effect of somatostatin is mediated via specific somatostatin receptors. Somatostatin does not inhibit SDF-1alpha-induced T cell attachment to the collagen substrate, which indicates that this neuropeptide specifically inhibits the process of chemokine-induced T cell penetration and migration through the collagen.

Abnormal expression of chemokine receptors on T-cells from patients with systemic lupus erythematosus.

The expression of chemokine receptors on T-cells and chemokine levels in the blood was studied in 23 patients with SLE (ACR criteria), seven patients with rheumatoid arthritis (RA) and in 15 healthy controls using flow cytometry, RT-PCR and ELISA. The cell surface expression of the chemokine receptors CXCR5 and CCR6 was decreased in SLE patients compared with controls (P = 0.051 and P = 0.002, respectively). The decrease of CXCR5 was confined to SLE patients with inactive disease (SLEDAI < 6) compared with active disease (SLEDAI > 6) and controls. CXCR2 and CCR1 were increased in patients with active SLE compared with patients with inactive disease (P = 0.001 and P = 0.01, respectively) and with controls (P = 0.02 and P = 0.053, respectively). The levels of the chemokines MIP-1alpha MCP-1, SDF-1alpha, IP-10 and RANTES were significantly elevated in SLE patients compared with controls. Patients with renal involvement had increased surface expression of CXCR3 and CCR3 (P = 0.04 in both) and a lower level of soluble IP-10 compared with patients without renal disease (P = 0.025) and compared with controls (P = 0.001). The ratio between CCR5 and CCR3 was significantly increased in RA patients compared with SLE patients and controls supporting a Th1 overweight in RA. In conclusion, patients with SLE showed abnormal T-cell expression of several chemokine receptors and levels of soluble chemokines in their plasma/serum.

Somatostatin receptor (SSTR) expression and function in normal and leukaemic T-cells. Evidence for selective effects on adhesion to extracellular matrix components via SSTR2 and/or 3.

We have examined normal T-cells and T-cell lines with respect to expression of various somatostatin receptor subtypes (SSTR1--5) using RT-PCR and PCR. To evaluate the function of these receptors we have further studied the effects of subtype specific signalling on T-cell adhesion using somatostatin analogs specific for various receptors as probes. Human T-lymphocytes showed SSTR expression related to activation and stage of differentiation. Normal T-cells (peripheral blood, T-cell clone) and T-leukaemia cell lines expressed SSTR2, SSTR3 and SSTR4. Normal T-cells expressed SSTR1 and SSTR5 while T-leukaemia lines did not. SSTR5 was selectively expressed in activated normal T-cells. T-lymphocytes produced no somatostatin themselves. Somatostatin and somatostatin analogs specific for SSTR2 and/or SSTR3 enhanced adhesion of T-cells to fibronectin (FN), and to a certain extent, also to collagen type IV (CIV) and laminin (LAM). T-lymphocytes express multiple SSTR and somatostatin may therefore regulate lymphocyte functions via distinct receptor subtypes as shown here for adhesion to extracellular matrix components (ECM) via SSTR2 and SSTR3. SSTR expression also distinguishes normal and leukaemic T-cells. Our findings suggest that SSTR subtypes may be useful targets for therapy during inflammatory diseases and malignancies affecting lymphocytes.

The regulation of FasL expression--a distinquishing feature between monocytes and T lymphocytes/NK cells with possible implications for SLE.

Monocytes and lymphocytes from patients with systemic lupus erythematosus (SLE) had a higher cell surface expression of FasL than the corresponding cells from healthy individuals. Inhibitors of metalloproteases upregulated the surface expression of FasL in peripheral blood lymphocytes (PBL), indicating that a metalloprotease is responsible for the cleavage of FasL. The level of sFasL in serum was slightly increased in the patient group compared to the controls. Therefore, the possible contribution of various mononuclear cell types to the release of FasL was analyzed. Isolated NK cells and T lymphocytes released FasL into the medium and the release was prevented by inhibitors of metalloproteases. In contrast, isolated monocytes did not release FasL. FasR expression was elevated in patients with inverted CD4/CD8 ratio, while FasL expression showed no relationship to CD4/CD8 ratio. The absence of FasL release by isolated cells and a high level of surface expression of FasL distinguish monocytes and T lymphocytes/NK cells.

Increased prevalence of atherosclerosis in patients with medium term rheumatoid arthritis.

OBJECTIVE: To measure the extent of atherosclerosis in patients with rheumatoid arthritis (RA) with a disease duration of considerable length, and in age and sex matched individuals. METHODS: Thirty-nine patients with RA (30 women, 9 men) with disease onset occurring between 1974 and 1978, and less than 65 years of age at the time of investigation, were enrolled together with 39 sex and age matched controls. Quantitative measurement of intima-media thickness (IMT) and semiquantitative assessment of the presence of plaque were undertaken by B-mode ultrasound of the common carotid artery (CCA-IMT) and the common femoral artery on the right-hand side. Echo Doppler cardiography was performed with an Accuson Aspen. The results were related to disease activity variables and accumulated disease activity, to lipid levels [i.e., cholesterol, high density lipoproteins, low density lipoproteins, triglycerides (TG)], to hemostatic factors [tissue plasminogen activator antigen (tPAag), plasminogen activator inhibitor-1 (PAI-1), von Willebrand factor (vWF)], and to soluble adhesion molecules (sICAM-1 and sE-selectin). RESULTS: Patients with RA had higher maximal and mean IMT values compared with controls. The difference concerning mean CCA-IMT reached statistical significance in patients with RA and correlated significantly with lipids (cholesterol, LDL, LDL/HDL ratio, TG) and tPAag. The prevalence of plaques, as well as of aortic cusp sclerosis, was higher in RA but only the difference in aortic cusp sclerosis was statistically significant. Patients with plaques had significantly higher levels of lipids (cholesterol, LDL, LDL/HDL ratio) than patients without plaques, while patients with cusp sclerosis had significantly higher cholesterol and TG levels. sICAM-1 was significantly higher both in patients with plaques and in those with aortic cusp sclerosis compared to patients without. CONCLUSION: Our results suggest an accelerated atherosclerosis in patients with RA that is related mainly to lipid levels.

Defective chemokine production in T-leukemia cell lines and its possible functional role.

Peripheral blood lymphocytes and T-cell clones produced nanogram quantities of the chemokines RANTES, MIP-1alpha, MIP-1beta, MCP-1, IL-8 and GRO-alpha as well as the motogenic cytokine HGF. In contrast, various T-leukemia cell lines at different stages of differentiation did not produce the same chemokines/cytokines. In order to study the possible functional importance of the poor chemokine production different T-cell lines were compared with respect to development of motile forms and migration on extracellular matrix components in the absence and presence of various chemokines. RANTES, MIP-1alpha, MIP-1beta, IL-8, GRO-alpha and lymphotactin did not augment the development of motile forms including the size and appearance of the pseudopodia activity of the T-leukemia cell lines. The T-cell lines migrated spontaneously on/to fibronectin in a Boyden chamber assay system. Chemokines augmented the migration of the T-leukemia cell lines on fibronectin in the Boyden system in a chemotactic fashion with peak responses at 10 to 50 ng/ml. Thus, the production of chemokines is defective in neoplastic T-lymphocytes. The defective chemokine production does not seem to play any major role for the basic locomotor capacity of the cells but may modulate the responsiveness to exogenous chemokines.

Cytocidal and apoptotic effects of the ClyA protein from Escherichia coli on primary and cultured monocytes and macrophages.

Cytolysin A (ClyA) is a newly discovered cytolytic protein of Escherichia coli K-12 that mediates a hemolytic phenotype. We show here that highly purified ClyA and ClyA-expressing E. coli were cytotoxic and apoptogenic to fresh as well as cultured human and murine monocytes/macrophages.

gammadelta T cells of human early pregnancy decidua: evidence for cytotoxic potency.

The immune compromise in decidua allows a semiallogeneic fetus to survive without impairing the ability of the maternal immune system to fight infections. Cytotoxic mechanisms are likely to be important in this compromise. Using RT-PCR, immunoflow cytometry and immunoelectron microscopy, the cytotoxic potential of isolated human decidual gammadelta T cells was studied. mRNA for perforin (Pf), granzymes A and B, granulysin and Fas ligand (FasL) was simultaneously expressed in decidual gammadelta T cells. Pf and FasL were not expressed on the cell surface. However, the cells constitutively synthesized Pf and stored it in cytolytic granules. Within the granules Pf mainly resided in the granule core formed by Pf-containing microvesicles. Ultrastructurally, three groups of Pf-containing granules were distinguished. They probably represent different stages of granule maturation in a process where Pf-containing microvesicles first attach to the core cortex and then are translocated across the cortex into the core. Presynthesized FasL was also stored in the core and microvesicles of the cytolytic granules. Upon degranulation by ionomycin/Ca(2+) treatment, FasL was rapidly translocated to the cell surface, demonstrating that its surface expression was not controlled by de novo biosynthesis. Thus decidual gammadelta T cells appear to perform Pf- and FasL-mediated cytotoxicity utilizing a common secretory mechanism based on cytolytic granule exocytosis. The first cytochemical visualization of lipids in the cytolytic granules is provided. These intragranular lipids probably wrap up the core and participate in packaging of the cytotoxic proteins as well as in the killing process. An ultrastructural model of a cytolytic granule is presented.

T cell activation in patients with ANCA-associated vasculitis: inefficient immune suppression by therapy.

BACKGROUND AND AIMS: Patients with vasculitic disease and autoantibodies to neutrophil cytoplasmic antigens (ANCA) generally respond to immunosuppressive therapy with a reduction of the inflammation and lowering of the ANCA titre. However, most patients experience relapses, sometimes after years of quiescence. In the present study we addressed the question whether the relapsing nature of this disease could be dependent on an underlying T cell activation. Patients were analyzed at disease onset, in remission while on treatment, and in quiescence. PATIENTS AND METHODS: Blood lymphocyte subsets and the expression of molecules associated with T cell activation were analyzed by flow cytometry and soluble interleukin-2 receptor (sIL2r) levels in serum by ELISA. Three patient categories (la, 1b and 2) were studied and compared with age-matched healthy controls (1a: 16 patients at onset of the disease before therapy, 1b: 10 patients from group 1a, re-analyzed after first remission, 2: 11 other patients in quiescence, 2-10 years after debut). RESULTS: All patient groups, 1a, 1b and 2, showed signs of T cell activation such as reduced CD28 on CD3+ and increased of the early T cell activation marker CD69 on CD3+, as well as of CD38 on CD8+ T cells. The sIL2r levels were significantly raised in all patient categories (la: 4280, 1b: 1844, 2: 2882 ng/ml) compared with the controls (923 ng/ml). CONCLUSION: Patients with ANCA-positive vasculitis show an increased expression of T cell activation markers irrespective of immunosuppressive therapy or disease phase. Such memory cells may form the basis for the remitting course of vasculitides and would be a rational target for new strategies of therapy.

Adhesion molecules (E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1)) in sera from patients with Staphylococcus aureus bacteraemia with or without endocarditis.

The aim of this prospective study was to evaluate if patients with endocarditis display a more extensive endothelial activation than those with bacteraemia but without endocarditis. Sixty-five patients with blood culture-verified Staphylococcus aureus bacteraemia were included and serum samples collected on admission were analysed by enzyme immunoassays. Elevated serum concentrations of adhesion molecules were found in most of the patients with S. aureus bacteraemia. Patients with endocarditis (n = 15) showed significantly higher serum E-selectin (median 156 ng/ml) and VCAM-1 (median 1745 ng/ml) concentrations compared with those with S. aureus bacteraemia but without endocarditis (80 ng/ml and 1172 ng/ml, respectively; P = 0.01 and P = 0.003). No significant difference was found between the groups concerning ICAM-1 (median 451 ng/ml versus 522 ng/ml). In addition, serum tumour necrosis factor-alpha (TNF-alpha) concentrations were significantly correlated (P < 0.002) to serum levels of E-selectin, ICAM-1 and VCAM-1.

Spectrum of extracellular matrix degrading enzymes in normal and malignant T lymphocytes.

Human T cells produce and release fibronectin degrading neutral serine proteases with a molecular weight of 50 kD, 70-80 kD (doublet) and 95 kD and have a cell associated 400 kD fibronectin degrading enzyme. In addition, human T cells produce proteases with m.w. 50, 70-80 kD and 400 kD which degrade laminin. CD 4+ T lymphocytes from a non-malignant cloned human T cell line produce a 92 kD gelatinase (MMP 9) and malignant T cell lines release, in addition to the 92 kD gelatinase, low amounts of a 72 kD gelatinase (MMP 2). Purification of the enzymatic activities using benzamidine sepharose yields a 50 kD and a 70 kD band of which the 50 kD band has fibronectin degrading capacity. The purified enzymes do not react with monoclonal antibodies to various previously characterized proteolytic enzymes present in T cells. T lymphocytes from a non-malignant cloned human T cell line produce high amounts of the 50 and 70-80 kD proteases directly after stimulation with anti-CD 3 monoclonal antibodies whereafter the production of these enzymes declines with time. The expression of the 400 kD fibronectin-degrading protease is downregulated by crosslinking of alpha 4 beta 1-integrin receptors on T cells using monoclonal antibodies. Thus, T lymphocytes produce several matrix degrading enzymes with multiple substrate specificities. The expression of these enzymes is controlled partly by lymphocyte activation signals or by direct signalling via beta 1-integrins.

Somatostatin- and factor XIIIa-immunoreactive cells in psoriasis during clobetasol propionate and calciprotriol treatment.

This study describes the changes in number and distribution of somatostatin- and factor XIIIa-immunoreactive dendritic cells in the epidermis and dermis of psoriatic lesional skin during topical treatment with clobetasol propionate or calcipotriol. Immunohistochemical analysis showed that the number of each cell type was increased in lesional skin as compared to normal skin. Investigation of serial biopsies from psoriasis lesions revealed a significant reduction in the number of somatostatin- and factor XIIIa-positive dendritic cells during the treatments. The reduction rate of the somatostatin-positive cells differed between the two groups and closely paralleled the healing process induced by the two treatments. These findings and the fact that somatostatin has been used in several studies as treatment for psoriasis may indicate that the somatostatin-positive cells are specifically involved in the healing process of psoriasis. The reduction of the factor XIIIa-positive cells was associated with the healing process as a whole, but showed no relation to either treatment.

Diminished interleukin-6 and C-reactive protein responses to laparoscopic versus open cholecystectomy.

BACKGROUND: Cytokines and their inhibitors are thought to be involved in many of the pathophysiological changes associated with trauma and infection. The magnitude of the trauma and the degree of tissue damage have an impact on the trauma response. The purpose of the study was to examine cytokine and hormonal responses to elective cholecystectomy and the extent to which these responses are influenced by the surgical procedure employed. METHODS: Sixteen patients, ASA grades I and II, were studied: 8 of them underwent laparoscopic cholecystectomy while the remaining 8 were operated on using the open technique. Systemic concentrations of tumour necrosis factor alpha (TNF), interleukin-1 beta (IL-1), interleukin-6 (IL-6), cortisol, epinephrine and norepinephrine were measured before and during the operation and subsequently for up to 48 h postoperatively. The degree of pain and fatigue were recorded during the study period. RESULTS: The preoperative levels of cytokines and hormones were all similar in the groups. Concentrations of TNF and IL-1 were detected only sporadically. The rise in plasma IL-6 was less marked following laparoscopic than after open cholecystectomy. However, the hormonal response was quite similar in the two groups. Pain and fatigue scores were lower (P < 0.05-0.01) in the laparoscopic group than in the open surgery group. CONCLUSION: In summary, cholecystectomy, irrespective of whether it was performed using the laparoscopic or open technique, was followed by a trauma response and increased pain and fatigue. However, the magnitude of stress, pain and fatigue was less pronounced in laparoscopic cholecystectomy patients. Concentrations of IL-6 seem to be more sensitive when it comes to delineating the trauma response than systemic norepinephrine and epinephrine levels.

Infiltrative capacity of T leukemia cell lines: a distinct functional property coupled to expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinases-1 (TIMP-1).

Infiltrative capacity was found to distinguish separate T leukemia cell lines. Of seven T-cell lines four exhibited capacity to infiltrate Matrigel. Analysis of infiltration was performed at the single-cell level throughout the Matrigel using a depth meter. Further, we examined differences in migration capacity and metalloproteinase production between infiltrating and non-infiltrating T-cell lines. The capacity to infiltrate was not directly correlated to the capacity to adhere to the Matrigel or to migrate on/to extracellular matrix components. It is concluded that infiltration capacity does not simply reflect capacity to migrate but represents a distinct functional property. The production of metalloproteinases and their inhibitors by the separate T-cell lines was analyzed using rt PCR, biosynthetic labelling, zymography, immunoprecipitation and ELISA. All T-cell lines with capacity to infiltrate produced matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) while non-infiltrating cell lines did not express MMP-9. Expression of MMP-1, 2, 3, 10, 14 and 17 showed no correlation to capacity to infiltrate. Analysis of infiltration in the presence of a metalloprotease inhibitor showed an increased number of cells within the gel. This enhancement of infiltration suggests that the function of MMPs and/or their inhibitors in lymphocyte infiltration is more complex than previously thought.

Cytokines and adhesion molecules in patients with polymyalgia rheumatica.

Serum levels of interleukin-1beta (IL-1beta), IL-1 receptor antagonist (IL-1ra), tumour necrosis factor alpha (TNF-alpha), IL-6, soluble IL-6 receptor (sIL-6R), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble E-selectin were measured in 15 patients with newly diagnosed polymyalgia rheumatica (PMR) before and after 3 months of corticosteroid therapy. Both IL-6 and IL-1ra were significantly increased in untreated PMR and remained elevated compared with controls during therapy, although significantly only for sIL-1ra. sICAM-1 was raised in 12/15 (87%) patients at diagnosis and remained high in 10/14 (71%) patients; soluble E-selectin levels were initially raised in 6/15 (40%) patients and decreased with therapy in those with the highest levels. IL-6, IL-1ra and sICAM-1 are sensitive indicators of continuing immunological activation in PMR; the advantages of these markers in assessing the response to therapy should be investigated in a longitudinal study.

In vitro Leishmania major promastigote-induced macrophage migration is modulated by sensory and autonomic neuropeptides.

Recruitment, migration and adherence of macrophages and their interaction with inoculated promastigotes are key steps in the initiation of the inflammatory process in cutaneous leishmaniasis. Parasite- and nervous system-derived factors might be involved in this process. In the present study the chemotactic activities of live, killed and sonicated Leishmania major promastigotes and of the promastigote culture supernatant as well as the L. major surface protease gp63 towards a murine macrophage cell line, Raw 264.7, were investigated, using the Boyden technique. The sensory neuropeptides SOM, CGRP and SP, and the autonomic neuropeptides VIP and NPY, were also investigated for possible modulatory effects on this chemotaxis, using the living promastigotes. Living promastigotes were the most efficient attractants for macrophages compared with other forms of the parasites. Prior incubation of the macrophages with the parasites completely abolished the chemotactic activity. This might indicate that the living promastigote chemotaxis is a receptor-mediated process. On the other hand, paraformaldehyde-killed promastigotes not only failed to induce macrophage chemotaxis but also inhibited it in comparison with the control. The surface protease gp63 tended to inhibit the macrophage chemotactic activity and the sonicate tended to stimulate it compared with controls. The culture supernatant had no effect, indicating that the chemoattractive factors putatively synthesized by the living promastigotes are not released to the surrounding medium. Somatostatin inhibited L. major promastigote-induced macrophage migration at a high concentration, 10(-6) M, while substance P inhibited it at both low concentrations, 10(-10) and 10(-9) M, and a high one, 10(-6) M, the last-mentioned having the greatest inhibitory effect. A stimulatory effect of calcitonin gene-related peptide was found at high concentrations, 10(-5) and 10(-6) M. Vasoactive intestinal peptide stimulated macrophage chemotactic activity at both a high, 10(-5) M, and at a low, 10(-9) M, concentration, the same concentration at which neuropeptide Y exerted its maximum inhibitory effect.

A tumor derived motility factor that stimulates cell migration on extracellular matrix.

A factor that stimulates migration of lung carcinoma cells on biological substrata was purified from the human lung adenocarcinoma cell line WART. A partially purified autocrine motility factor-like substance, termed haptotaxin, was added to the lower compartment of Boyden chambers and the filters were coated on the upper, lower or both sides with different concentrations of the extracellular matrix (ECM) components fibronectin, laminin or collagen type IV. These adhesive proteins coated on the lower surface of the filter promoted the migration (haptotaxis) of lung carcinoma cells. This effect was greatly enhanced by the addition of haptotaxin. In contrast, ECM components (including gelatin) coated on the upper surface or on both filter surfaces did not stimulate tumor cell migration. However, the addition of haptotaxin also timulated cell migration under these conditions. Haptotaxin did not stimulate migration on filters coated with bovine serum albumin or on uncoated filters. Haptotaxin could not be absorbed by fibronectin, laminin, collagen type IV or gelatin, and soluble ECM components did not affect the locomotor effect of haptotaxin. Substrata coated with fibronectin, laminin and collagen type IV induced adhesion and spreading of lung carcinoma cells in a dose dependent fashion. Haptotaxin potentiated adhesion and spreading of tumor cells on these substrata but did not in itself mediate adhesion and spreading of the cells. Anti-VLA 2 antibodies inhibited migration to haptotaxin on gelatin and laminin coated filters but did not affect haptotaxin-induced migration on fibronectin or collagen type IV substrata. Anti-VLA-5 monoclonal antibodies inhibited haptotaxin-induced migration on fibronectin coated filters but not such migration on filters coated with other ECM molecules showing that tumor cells must interact specifically with ECM components in order to migrate to haptotaxin.