E L Filonzi

Cytokine regulation of granulocyte-macrophage colony stimulating factor and macrophage colony-stimulating factor production in human arterial smooth muscle cells

Smooth muscle cells (SMC) are the major cell type found in the walls of large blood vessels and appear to participate in local immune and inflammatory reactions, as well as in certain vascular diseases. We tested whether human arterial SMC can produce in vitro the colony stimulating factors (CSFs), granulocyte macrophage-CSF (GM-CSF) and macrophage CSF (M-CSF). Untreated internal mammary artery and aortic SMC produced no detectable GM-CSF but constitutively made M-CSF, measured by ELISA and radioimmunoassay, respectively. Interleukin-1 (IL-1) and, to a lesser extent, tumor necrosis factor alpha (TNF alpha) stimulated GM-CSF formation within 3 h; mRNA levels also increased particularly in the presence of the protein synthesis inhibitor, cycloheximide. IL-1, TNF alpha and, in addition, interferon-gamma (IFN-gamma) raised the M-CSF levels within 6 h; cycloheximide potentiated the effects of IL-1 and TNF alpha on mRNA levels. These results suggest that cytokine-stimulated human arterial SMC may be a source of the M-CSF found in atherosclerotic lesions. Since monocytes/macrophages can be activated by GM-CSF and M-CSF, while GM-CSF can also affect granulocyte function, SMC may participate in inflammatory reactions and vascular diseases by releasing these cytokines.

Regulation of Macrophage Colony-Stimulating Factor (M-CSF) Production in Cultured Human Synovial Fibroblasts

OBJECTIVE To study the regulation of macrophage-colony stimulating factor (M-CSF) formation in vitro by human synovial fibroblast-like cells. METHODS Human synovial cell explant cultures were established using cells from non-rheumatoid donors. M-CSF antigen was measured by immunoassay, and messenger RNA (mRNA) levels were determined by Northern blot. RESULTS The cytokines, interleukin-1 (IL-1), tumor necrosis factor alpha (TNF alpha), interferon-gamma (IFN-gamma) and IL-4, increased production of M-CSF above constitutive levels. The presence of the cyclooxygenase inhibitor, indomethacin, potentiated the action of IL-1 on M-CSF synthesis, suggesting that an endogenous cyclooxygenase product(s) can down-regulate M-CSF formation. Changes in M-CSF mRNA levels paralleled those in protein levels. The glucocorticoid, dexamethasone, and the retinoid, all-trans retinoic acid, stimulated M-CSF formation. The control of M-CSF synthesis in the synovial fibroblasts differs from that for granulocyte macrophage-CSF (GM-CSF) and granulocyte-CSF (G-CSF). CONCLUSION These results suggest that cytokine-stimulated synovial fibroblasts may be a source of M-CSF production in the joints of patients with inflammatory arthritis; as a result, monocyte/macrophages may be activated, leading to perpetuation of the inflammation and destructive events occurring in these lesions.

Stimulation of PAI-1 expression in endothelial cells by cultured vascular smooth muscle cells.

Regulation of endothelial cell (EC) plasminogen activator inhibitor type-1 (PAI-1), the primary physiological inhibitor of tissue-type plasminogen activator (TPA) and urokinase-type plasminogen activator (UPA), by various stimuli has been well characterized. We report the upregulation of secreted and intracellular PAI-1 in human umbilical ECs when cocultured with human smooth muscle cells (SMCs) on amniotic membranes or incubated with SMC conditioned medium (CM) under serum-free conditions as determined by enzyme-linked immunosorbent assay. Cocultured human umbilical vein ECs and SMCs, or human umbilical artery ECs and SMCs, displayed a 73% and 68% increase, respectively, in released PAI-1. SMC-derived stimulatory factor release showed tissue specificity, since only human aortic, umbilical vein, and umbilical artery SMCs upregulated PAI-1 synthesis, whereas SMCs from human mammary artery, pulmonary artery, and saphenous vein did not. Stimulation of EC PAI-1 by SMC CM was both time and concentration dependent, with as much as five- and fourfold increases in supernatants and lysates, respectively. PAI-1 synthesis and activity in ECs from other vascular beds were also upregulated by SMC CM. Northern blot analysis paralleled the protein results, showing as much as a 2.7-fold increase in specific EC PAI-1 mRNA expression after incubation with SMC CM for 8 hours. PAI-1 stimulatory activity in SMC CM was completely abolished by boiling or incubation with protamine sulfate and was reduced by transient acidification or heparin-Sepharose pretreatment by 33% or 48%, respectively. The stimulatory factor(s) appeared to have a molecular mass of 23 kD as determined by gel filtration.(ABSTRACT TRUNCATED AT 250 WORDS)