Tumor necrosis factor alpha stimulates production of leukemia inhibitory factor in human dermal fibroblast cultures.

Leukemia inhibitory factor (LIF) is a recently described cytokine with a variety of actions including a possible involvement in immune responses. We determined whether human dermal fibroblast cultures could produce LIF after they were treated with tumor necrosis factor alpha (TNF alpha), a cytokine that is produced as an early inflammatory response of activated monocytes. We found that treatment of the cultures with as little as 0.5 units/ml (1.5 pM) caused a detectable increase in both LIF message and protein as measured by Northern blot assay and ELISA, respectively. Furthermore, increasing concentrations of TNF alpha produced a dose-dependent increase in both steady-state LIF mRNA and protein levels up to a maximum response with 500 units/ml (1.5 nM). Increases in LIF mRNA levels were rapid and could be detected 1 hr after treatment with 500 units/ml of TNF alpha. However, this effect was transient. It reached a maximum at 2 hr and returned almost to baseline at 24 hr. In contrast, levels of LIF protein in the conditioned media of the cultures increased progressively over 24 hr. The LIF produced by these cultures was biologically active and was inhibited by a polyclonal antibody to human LIF in a bioassay. These results demonstrate that LIF is produced by human dermal fibroblasts in response to treatment with TNF alpha, a mediator of acute inflammation. Furthermore, they suggest that production of LIF by these cells may be involved in the development of both the local and generalized immune response.

Production of leukemia inhibitory factor mRNA and protein by malignant and immortalized bone cells.

Leukemia inhibitory factor (LIF) is a recently characterized glycoprotein with complex biologic activities on bone cells. We tested various rodent and human immortalized and malignant bone cell lines and primary osteoblast-enriched cell cultures from fetal rat calvarial digests for expression of LIF mRNA and LIF protein. Both human and rodent immortalized and malignant cells expressed a single 4.4 kb mRNA transcript that hybridized to a human LIF cDNA probe in Northern blots. LIF mRNA was undetectable in unstimulated rodent osteoblast-like cells lines MC3T3-E1 and Py1a. However, treatment with LPS (10 micrograms/ml), TGF-beta (1 ng/ml), TNF-alpha (100 ng/ml) or inhibitors of protein synthesis (cycloheximide, emetine, puromycin, and anisomycin) induced the expression of LIF message in these cells. In contrast, primary osteoblast-enriched cells did not express LIF mRNA in Northern blot assays either constitutively or after treatment with TNF-alpha or cycloheximide. The human osteosarcoma cells lines U-2 OS and Saos-2 constitutively expressed LIF mRNA and did not respond to LPS treatment. However, phorbol myristate acetate (PMA), an activator of protein kinase C, was a potent stimulator of LIF message in Saos-2 but not U-2 OS cells. The effects of PMA (0.5 ng/ml) on LIF mRNA in Saos-2 cells were detectable at 1 h and maximal at 6 h. TNF-alpha (100 ng/ml) and inhibitors of protein synthesis also increased LIF mRNA in both Saos-2 and U-2 OS cells. LIF protein was also detected constitutively in the conditioned medium from both Saos and U-2 OS cells.(ABSTRACT TRUNCATED AT 250 WORDS)