Niclas Olsson

Human mast cell migration in response to members of the transforming growth factor-beta family.

Mast cells are known to accumulate at sites of inflammation, however, the chemotaxins involved remain largely undefined. Transforming growth factor‐β (TGF‐β) isoforms regulate numerous cellular functions, including cell growth and differentiation, formation of extracellular matrix, and the immune response. In this study we have compared the potency of different members of the TGF‐β family as human mast cell chemotaxins, and analyzed the expression of TGF‐β binding proteins on human mast cells. We were able to demonstrate that the maximal chemotactic response was attained at approximately 40 fM for the three TGF‐β isoforms, with TGF‐β3 being more effective than TGF‐β1 and TGF‐β2 at this concentration. This effect was observed in both the HMC‐1 human mast cell line and in cultured primary mast cells. In addition, TGF‐β1, TGF‐β2, and less efficiently, TGF‐β3 inhibited the proliferation of HMC‐1 cells. The migratory response is probably mediated through interaction with the TGF‐β serine/threonine type I and II receptors that were found to be expressed on the cells. No expression of TGF‐β type III receptor, endoglin, or the endothelial TGF‐β type I receptor ALK‐1 could be detected. These results provide evidence that TGF‐β isoforms are highly potent chemotaxins for human mast cells and can play an important role in the recruitment of mast cells in inflammatory reactions. J. Leukoc. Biol. 67: 350–356; 2000.

Expression of CCL5/RANTES by Hodgkin and Reed-Sternberg cells and its possible role in the recruitment of mast cells into lymphomatous tissue

HL is a malignant lymphoma characterized by a small number of malignant HRS cells among a major population of infiltrating reactive cells, e.g., lymphocytes and eosinophils. We previously reported that mast cells are present in HL‐affected lymph nodes and therein are the predominant CD30L‐expressing cells. The CD30L expressed on mast cells is functionally active and can provide stimulatory signals to HRS cells. Thus, mast cells constitute an important portion of the infiltrating reactive cells that contribute to tumor progression in HL. Control of the recruitment of this previously unrecognized cell and its interactions with tumor cells are essentially unknown. To elucidate if mast cells might be specifically attracted to the tumor area by chemokines produced by HRS cells, we investigated chemokine expression in HL cell lines and in vivo. By RNase protection assay, mRNA expression of several chemokines could be detected in the cell lines. Despite the heterogeneous expression profile exhibited by the cell lines, 4 of 5 expressed CCL5 (RANTES) mRNA. RT‐PCR and immunohistochemistry confirmed expression of CCL5 in vivo. Furthermore, secreted CCL5 was detected in conditioned media from 3 of the cell lines. In a migration assay, we found that CCL5 present in conditioned medium could induce mast cell migration. Taken together, our results suggest that CCL5 produced by HRS cells is one mechanism by which mast cells can be attracted into the tumor tissue in HL.

The chemokine receptor CXCR4 is expressed within the mast cell lineage and its ligand stromal cell‐derived factor‐1α acts as a mast cell chemotaxin

In this study we provide evidence that the chemokine stromal cell‐derived factor‐1α (SDF‐1α) acts as a mast cell chemoattractant through interactions with its receptor CXCR4 expressed on mast cell progenitors in the blood as well as on in vitro‐developed and leukemic mast cells. We found expression of CXCR4 on cord blood‐derived mast cells (CBMC) and on the human mast cell line HMC‐1, analyzed by RNAse protection assay and flow cytometry. SDF‐1α induced intracellular calcium mobilization in HMC‐1 cells and was chemotactic for both HMC‐1 cells and CBMC. The activity of SDF‐1α was completely blocked by treating the cells with pertussis toxin, indicating the involvement of Gi‐proteins in the signaling. By applying a transwell assay we could show that SDF‐1α induces migration of a cell population in peripheral blood that is enriched for cells with the capacity to differentiate into mast cells. These findings thus suggest a mechanism by which human mast cell progenitors may be recruited from circulation into the tissue.

Selective CCL5/RANTES-induced mast cell migration through interactions with chemokine receptors CCR1 and CCR4

Mast cells (MCs) accumulate at sites of allergic mucosal inflammation where they act as central effector and regulatory cells. Because chemokines are of vital importance in directing inflammatory leukocytes to the sites of inflammations, we have investigated the expression and function of CC-chemokine receptor (CCR) on human MCs. Two previously unrecognized MC-chemokine receptors, CCR1 and CCR4, could be identified on cord blood-derived MCs (CBMCs). CCR1 and CCR4 expressed on CBMCs exhibited a unique response profile. Of seven CCR1 and CCR4 agonists tested, only CCL5/RANTES act as an agonist inducing chemotaxis. The migration could be partially blocked by specific antibodies against CCR1 or CCR4, while a complete inhibition was achieved when both CCR1 and CCR4 were blocked. These results demonstrate that both CCR1 and CCR4 are functional receptors on human mast cells with capacity to mediate migration towards CCL5.

Transforming growth factor-β-mediated mast cell migration depends on mitogen-activated protein kinase activity

Abstract Transforming growth factor-β (TGF-β) isoforms regulate numerous cellular functions through binding to receptors with intrinsic serine/threonine kinase activity that transduce the intracellular signals via activation of Smad proteins. In this study, we examined the signalling pathways involved in TGF-β1-mediated growth inhibition and migration in a human mast cell line, HMC-1. TGF-β1 evoked optimal migration at 40 fM, whereas maximal growth inhibition was obtained at 400 pM. Protein tyrosine kinase inhibitors completely inhibited TGF-β1-mediated migration, without affecting the antimitogenic response. Smad2 was phosphorylated upon TGF-β1 treatment, both in the absence and presence of genistein. The mitogen-induced extracellular kinase (MEK) inhibitor, PD98059, blocked the migratory response without affecting growth inhibition. In contrast, the p38 MAP kinase inhibitor, SB203580, had no significant effect on either migration or growth inhibition. These results indicate that different signalling pathways mediate TGF-β1-induced migration and growth inhibition in HMC-1 cells, where the migration involves MEK activity.

Demonstration of mast cell chemotactic activity in synovial fluid from rheumatoid patients

OBJECTIVES The significance of the mast cell in the pathogenesis of rheumatic diseases has become more evident. Although mast cell hyperplasia is a feature of rheumatoid arthritis, the nature of mast cell chemoattractants involved in the recruitment of mast cells in joint diseases has not been studied in any detail. In this study the presence of mast cell chemotactic activity in synovial fluids was examined. METHODS Synovial fluids from seven rheumatoid patients were tested in a modified Boyden chamber, where a human mast cell line was used as responder. The presence of stem cell factor (SCF) and transforming growth factor β (TGFβ) was measured by enzyme linked immunosorbent assay (ELISA). RESULTS Six of the seven synovial fluids tested exhibited mast cell chemotactic activity. Two characterised human mast cell chemotaxins, SCF and TGFβ, were highly expressed in the synovium. Soluble SCF could be detected in all fluids analysed. Blocking antibodies against SCF or TGFβ almost completely blocked the activity in one fluid, partially blocked the activity in three, and did not affect the activity in two. Treatment of the responder cells with pertussis toxin reduced the migratory response against seven fluids, indicating the presence of chemoattractants mediating their effect through Gi coupled receptors. CONCLUSION These data demonstrate the presence of multiple factors in synovial fluid acting as mast cell chemoattractants, two of which are SCF and TGFβ that contribute to the effect. These findings may be of importance for developing new strategies to inhibit mast cell accumulation in rheumatic diseases.

Regulation of Mast Cell Migration by TH1 and TH2 Cytokines: Identification of Tumour Necrosis Factor‐α and Interleukin‐4 as Mast Cell Chemotaxins

Mast cells act as central effector and regulatory cells in many inflammatory disorders, including T helper 1 (TH1)‐mediated inflammations such as autoimmunity and TH2‐mediated inflammations such as allergy and parasite infections. One characteristic for mast cell‐mediated inflammations is the accumulation of mast cells in the inflamed tissue. The factors regulating mast cell recruitment in these inflammations are still not fully characterized. We have investigated the potency of TH1‐ and TH2‐secreted cytokines to mediate mast cell migration. Supernatants from six different TH1 and TH2 clones were tested for mast cell‐chemotactic activity using the human mast cell line (HMC‐1) as a responder cell. All six clones produced factors that induced mast cell migration. Using blocking antibodies to a broad range of cytokines, we found that anti‐tumour necrosis factor‐α (anti‐TNF‐α) reduced the migration of mast cells to supernatants from TH1 clones. In contrast, the main mast cell chemoattractants secreted by TH2 clones were found to be interleukin‐4 (IL‐4) and IL‐8. The potency of these cytokines to act as mast cell chemoattractants was confirmed by using recombinant IL‐4, IL‐8 and TNF‐α. Our results suggest that TNF‐α can be involved in the recruitment of mast cells in TH1‐mediated inflammations, whereas IL‐4 and IL‐8 might play a similar role in TH2‐mediated inflammations.

CCL17 and CCL22 attenuate CCL5‐induced mast cell migration

Background Mast cells (MCs) accumulate at sites of allergic mucosal inflammation where they act as central effectors and regulatory cells. Chemokines are believed to be crucial for the recruitment of MCs to sites of inflammation. We recently reported that human umbilical cord blood MCs (CBMCs) expresses the CC chemokine receptors, CCR1 and CCR4. We found a unique response profile to ligands of the respective receptors in which, of all tested ligands, only CCL5/RANTES‐induced migration.