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We have previously demonstrated that human promyelocytic HL-60 cells express transforming growth factor-alpha (TGF-alpha) during granulocytic differentiation. The present experiments were carried out in order to determine whether cells differentiated towards monocytes/macrophages will analogously express the TGF-alpha proto-oncogene product. HL-60 cells were induced to differentiate with 1 microM 1,alpha 25-dihydroxycholecalciferol (vitamin D3), and the human monocytoid cell line, U-937, was induced with 1 microM retinoic acid (RA), 0.1 microM vitamin D3, or 0.16 microM phorbol-12-myristate-13-acetate (PMA), ie experimental protocols known to induce monocyte/macrophage differentiation in these cells. In HL-60 cells, lacking constitutive TGF-alpha mRNA, vitamin D3 caused expression of the TGF-alpha gene and protein as demonstrated by Northern blot analysis and enzyme-linked immunoabsorbant assay (ELISA). In U-937 cells, showing constitutive TGF-alpha expression, RA but not vitamin D3 or PMA, caused marked increase in TGF-alpha mRNA (approximately 5-fold) and protein (approximately 3-fold) levels. In both cell lines the increase in TGF-alpha mRNA was evident within 24 h and continued throughout the observation period. Thus, it is established that differentiation of human leukemia cells towards monocytes/macrophages may be accompanied by TGF-alpha gene and protein expression in vitro. This is in conformity with the observed ability of mature activated macrophages to produce TGF-alpha.

We have previously demonstrated a constitutive expression of transforming growth factor alpha (TGF-alpha) in normal human blood eosinophils, both at the mRNA and protein level. This may indicate a novel function of the eosinophil, the regulation of which has not been clarified. Therefore human white blood cells (WBC) were treated with potential regulators of eosinophil function. Northern blot analysis demonstrated that human recombinant granulocyte-macrophage colony-stimulating factor (GM-CSF) or interleukin-3 (IL-3) caused a time and dose-dependent 2- to 3-fold increase of TGF-alpha mRNA levels, in relation to incubation in the absence of cytokine; maximal response was attained within 4 h of incubation. In contrast, IL-5 failed to influence the expression of the TGF-alpha gene. In situ analysis of GM-CSF- or IL-3-stimulated cells showed that eosinophils remained the sole cell type expressing TGF-alpha mRNA. However, whereas GM-CSF significantly induced, within 1 h, release of immunoreactive TGF-alpha protein, IL-3 was insufficient in this respect. In conclusion, our findings indicate that expression of TGF-alpha gene and protein in normal blood eosinophils is differently regulated by GM-CSF and IL-3.

Transforming growth factor alpha (TGF-alpha) is a pleiotropic factor mediating numerous cellular responses in normal and transformed cells. This includes differentiation, proliferation, migration, and formation of extracellular matrix. TGF-alpha has been demonstrated in circulating eosinophils from the idiopathic hypereosinophilic syndrome and in differentiating promyelocytic leukemia cells in vitro. Whether TGF-alpha production also occurs in normal human blood cells is not known. Northern blot analysis showed that normal human white blood cells consistently expressed the TGF-alpha gene in 47 out of 47 donors. Cell preparations enriched in mononucleated cells, and devoid of granulocytes, showed no TGF-alpha mRNA. In situ hybridization experiments assigned the TGF-alpha gene expression to the eosinophils; 100% of the eosinophils and no other cell types were specifically recognized by the complementary human TGF-alpha riboprobe. White blood cells, incubated at 37 degrees C for up to 6 hours, released immunoreactive TGF-alpha to the incubation medium, as determined by ELISA. In contrast, no TGF-alpha protein was detected in the incubation medium of mononuclear cells. It is concluded that TGF-alpha is constitutively produced and released by normal human blood eosinophils. TGF-alpha provided by eosinophils, may participate in the inflammatory reaction by interacting with mesenchymal and epithelial cells, thus promoting fibrosis or neovascularization.