Murine M290 integrin expression modulated by mast cell activation.

Intraepithelial T lymphocytes possess a (TGF beta)-inducible integrin complex that consists of the beta 7 integrin chain in association with a novel alpha chain, murine alpha M290. Using antibodies directed against the murine alpha M290 chain, IL-3-dependent murine bone marrow-derived mast cells demonstrated positive staining after induction with TGF beta. Sequence information from the human intraepithelial T lymphocyte-alpha E gene sequence was used to clone the murine homolog from a TGF beta-induced T cell library. The murine alpha E homolog gene encodes a protein recognized by antibodies to alpha M290. Expression of this gene by bone marrow-derived mast cells was found to be induced by TGF beta or IgE-mediated cross-linking of Fc epsilon R1. Furthermore, exposure of murine TK-1 cells to supernatants from activated mast cells induces the expression of the alpha M290 gene, indicating that a mast cell mediator(s) can act in a paracrine manner to induce expression of this integrin complex by other cells.

Preferential expression of interleukin-12 or interleukin-4 by murine bone marrow mast cells derived in mast cell growth factor or interleukin-3.

In this report we demonstrate that murine bone marrow cells cultured in either interleukin (IL)-3 or mast cell growth factor (MGF, also known as c-kit ligand and stem cell factor) differentially express cytokine genes. Bone marrow cells cultured in IL-3 differentiate and proliferate, taking on a mucosal mast cell-like phenotype. These cells express the IL-4 gene. Bone marrow cells cultured in MGF take on a connective tissue mast cell-like phenotype and possess transcripts for both of the subunits of the IL-12 cytokine. Bone marrow cells cultured in both IL-3 and MGF express the IL-4 gene at lower levels than that seen for the IL-3 culture alone, but do not possess IL-12 gene transcripts. The level of IL-12 subunit transcripts derived from the MGF-derived bone marrow cells was compared to that found in splenocytes and activated macrophages, the only cells in which IL-12 production has been previously documented. Both of the IL-12 subunit transcripts were found, compared to a beta-actin control, to be present within MGF-derived cells in the same if not higher quantities than the splenocyte or macrophage cultures. Mucosal mast cells have been previously implicated in the development of the T helper type 2 (TH2) T cell phenotype via their expression of IL-4. The finding that the MGF-derived connective tissue-like mast cells possess IL-12 transcripts suggests that the development of the TH1 T cell pathway may be positively influenced by this type of mast cell.

Modulation of integrin expression during mast cell differentiation.

Previously we have reported that rodent mast cells synthesize the mRNA encoding the alpha and beta integrin chains (alpha 4, beta 1 and beta 7) of the lymphocyte Peyer's patch adhesion molecule (LPAM)-1 and LPAM-2 lymphocyte homing receptors, and that they possess an alpha 4-containing integrin complex on their cell surface. In this report, we have examined the expression of these integrin chain genes by mature connective tissue mast cells (CTMC) and by bone marrow-derived mast cells (BMMC) differentiated from bone marrow precursor cells in the presence of interleukin (IL)-3 and/or the c-kit ligand (also known as mast cell growth factor and stem cell growth factor). High levels of both the beta 7 and beta 1 transcripts were present in mature CTMC while those encoding the alpha 4 chain were absent. Similarly, when BMMC were grown in IL-3 for 28 days and analyzed for integrin chain transcripts, those specific for the alpha 4 chain were also diminished compared to beta 7 and beta 1 transcripts. To compare the expression of these integrin genes during mucosal mast cell and CTMC development, BMMC were derived in the presence of IL-3 alone, c-kit alone, or IL-3/c-kit together. These experiments indicated that c-kit inhibited the transcription of the beta 7 and Fc epsilon RI genes while enhancing alpha 4 transcript levels. The enhancement of alpha 4 levels, however, was abrogated with the addition of IL-3. Similarly, the c-kit-induced depression of beta 7 and Fc epsilon RI transcript levels was overcome by the addition of IL-3. These data suggest that the integrin complexes synthesized by the mast cells may differ depending upon their path of differentiation and that another alpha chain integrin may be synthesized to complex with the beta 7 and/or beta 1 chains.

Expression of murine beta 7, alpha 4, and beta 1 integrin genes by rodent mast cells.

The screening of a rat mast cell cDNA library with a probe selected to recognize those genes preferentially or exclusively expressed by mast cells identified a rat gene sequence, RF-17, that shared homology with the beta-integrins. This integrin was expressed in rat tissues enriched for mast cells and T cells. The rat RF-17 sequence was used to isolate the murine homologue from a spleen cDNA library. The murine gene encodes a protein of 806 amino acids that is the probable homologue to the human beta 7 chain. Transcripts specific for the murine gene are found in the thymus, spleen, and lung. To attempt to identify the gene product for this new integrin chain, we examined the murine T cell line TK-1, which expresses a novel integrin heterodimer, lymphocyte Peyer's patch high endothelial venule adhesion molecule (LPAM-1), of a known alpha 4 chain and an unknown beta P chain, for expression of murine beta 7 (RF-17). This cell line expresses high levels of RF-17 transcripts, suggesting that beta P is encoded by the beta 7 gene. Bone marrow cells induced to differentiate into mast cells via IL-3 express the beta 7 gene as well as the genes encoding the murine integrin alpha 4 and beta 1 proteins. Surface staining analysis indicates that these cells express an alpha 4-containing integrin complex throughout the differentiation process. These data suggest that the Peyer's patch homing LPAM-1 receptor expressed by a subset of T cells consists of the beta 7 gene product and the alpha 4 chain, and that this integrin chain complex is also found on the surface of maturing mast cells. The presence of beta 1 transcripts also suggests that these maturing mast cells possess the LPAM-2 integrin complex (alpha 4/beta 1) as well. The experimental strategy described in this manuscript has, thus, identified a novel murine beta-integrin chain that is expressed by rodent T cells and mast cells.