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Transforming growth factor-b 1 (TGF-b 1) produced by in ltrating macrophage s plays a role in brotic disorders through the induction of myobroblasts. To explore possible mechanisms by which TGF-b 1 may act in this context, we investigated effects of TGF-b 1 on macrophage-like (HS-P) and myo broblastic (MT-9) cells, two novel cell lines developed by us. Immunocytochemicall y, the addition of TGF-b 1 (0, 0.1, 0.5, and 1.0 ng/ml) dose-dependentl y suppressed the expressions of antigens recognized by macrophage /histiocyte-speci c antibodies (ED1 and ED2) in HS-P cells, whereas the addition concomitantly increased the number of anti-a -smooth muscle actin antibody-positive myo broblastic cells, suggesting a possible phenotypica l modulation of macrophages into myo broblasts in the brotic lesions. By contrast, MT-9 cells did not show such immunophenotypica l changes following TGF-b 1 addition. DNA synthesis, measured by tritiated thymidine-incorporation , was inhibited in a dose-dependen t manner in MT-9 cells by TGF-b 1 addition (0, 0.1, 0.2, 0.5, 1.0, 5, and 10 ng/ml), but that in HS-P cells was unchanged . Northern blot analysis revealed that expressions of cell cycle-related early genes, c-jun and c-myc, were increased in HS-P cells after TGF-b 1 (1 ng/ml) addition, with c-jun showing peak expression prior to c-myc. By contrast, the peak expressions of c-jun and c-myc were delayed in TGF-b 1 (1 ng/ml)-added MT-9 cells, and their levels were less in MT-9 cells than in HS-P cells. Furthermore, TGF-b 1 (1 and 10 ng/ml) induced DNA laddering in MT-9 cells, but did not in HS-P cells. Based on these ndings, it was speculated that TGF-b 1 could have induced G1 arrest in cell cycle and apoptosis in MT-9 cells. The present study showed that there were signi cant differences in the effects of TGF-b 1 between macrophage-like HS-P cells and myo broblastic MT-9 cells, presumably depending on divergent susceptibilities to TGF-b 1 between both cell types. Because such cell types are key cells in the brogenesis, HS-P and MT-9 might be useful models for investigating the pathogenesi s of brosis in vitro.

Cellular intoxification by exotoxin A of Pseudomonas aeruginosa (PEA) begins when PEA binds to its cellular receptor, the low-density lipoprotein receptor-related protein (LRP). This receptor is particularly abundant on macrophages. We hypothesize here that inducible changes in cellular expression levels of the LRP represent an important mechanism by which macrophage susceptibility to PEA is regulated by the host. We have examined the effect of lipopolysaccharide (LPS) on LRP expression and PEA sensitivity in the macrophage-like cell line HS-P. Using a [3H]leucine incorporation assay to measure inhibition of protein synthesis, we have demonstrated that HS-P macrophages are highly sensitive to PEA and that PEA toxicity is decreased by the LRP antagonist receptor-associated protein. LPS pretreatment decreases HS-P PEA sensitivity in a time- and dose-dependent manner. The dose of toxin required to inhibit protein synthesis by 50% increased from 11.3 ± 1.2 ng/ml in untreated cells to 25.7 ± 2.0 ng/ml in cells treated with LPS. In pulse experiments, involving brief exposure to saturating concentrations of PEA, [3H]leucine incorporation was more than threefold higher in cells pretreated with LPS than in untreated macrophages. These changes in HS-P PEA sensitivity following LPS treatment were consistently associated with a fivefold decrease in HS-P LRP mRNA expression as measured by Northern blot analysis and a three-and-a-half-fold decrease in HS-P LRP-specific ligand internalization as determined by activated α2-macroglobulin internalization studies. These data demonstrate for the first time that modulation of LRP levels by extracellular signaling molecules can alter cellular PEA sensitivity.

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