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Published on: February 21, 2018
Differentiation-dependent modulation of TNF production by PAF in human HL-60 myeloid leukemia cells
M Rola-Pleszczynski1, J Stankova
1Department of Pediatrics, Faculty of Medicine, University of Sherbrooke, Québec, Canada.
Abstract:
Platelet-activating factor (PAF) can augment tumor necrosis factor (TNF) production by human monocytes in a bimodal manner, with two peaks of activation at picomolar and micromolar concentrations. These peaks are partially associated with monocyte subsets presenting different characteristics in terms of size, density, phenotypic markers, and [Ca2+]i mobilization responses. In the present study, we used the human promyelocytic leukemia cell line HL-60, at various times during differentiation with 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] toward the monocyte lineage, in order to study the relation of cell differentiation to responsiveness to PAF in terms of cytokine production. TNF production was induced by pretreatment with interferon gamma for 24 h and treatment with muramyl dipeptide. Although detectable TNF was produced by 4 day-differentiated cells, no effect was seen with PAF (10(-16)-10(-6) M) at this or earlier stages. In contrast, 5 day-differentiated cells had a comparable baseline production of TNF but responded with a 2.5-fold increase to PAF with a single peak, maximal at 10(-8) M. Moreover, 6 day- or 7 day-differentiated HL-60 cells showed a further increase in TNF production in response to PAF, and the response was bimodal, similar to that of the less dense subset of monocytes, with peaks at 10(-14) and 10(-7) M PAF. In parallel, undifferentiated HL-60 failed to respond to PAF in terms of [Ca2+]i mobilization. The earliest responsiveness to PAF (10(-7) M) was observed by 4 days of treatment with 1,25(OH)2D3, and by day 7 the response to PAF became bimodal (10(-14) and 10(-7) M). These results indicate that myeloid cells acquire, during maturation toward the monocyte lineage, a progressive responsiveness to PAF in terms of [Ca2+]i mobilization and enhanced cytokine production, and they suggest that the heterogeneity in responses to PAF observed in normal monocytes may be related to their stage of differentiation or maturation.
Insights
Platelet-activating factor (PAF) response in human monocytes increases with cell differentiation. Maturing myeloid cells show progressive calcium mobilization and enhanced tumor necrosis factor (TNF) production, explaining monocyte response variations.
Area of Science:
- Immunology
- Cell Biology
- Biochemistry
Background:
- Platelet-activating factor (PAF) modulates human monocyte cytokine production, including tumor necrosis factor (TNF).
- Monocyte responses to PAF are bimodal, with varying sensitivity linked to subset characteristics.
- The relationship between myeloid cell differentiation and PAF responsiveness requires further elucidation.
Purpose of the Study:
- To investigate how HL-60 cell differentiation, induced by 1,25-dihydroxyvitamin D3 [1,25(OH)2D3], affects responsiveness to Platelet-activating factor (PAF) in terms of cytokine production.
- To correlate the stage of myeloid cell maturation with calcium ion ([Ca2+]i) mobilization and TNF production in response to PAF.
Main Methods:
- Utilized the human promyelocytic leukemia cell line HL-60 undergoing differentiation towards the monocyte lineage.
- Administered 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] for varying durations (up to 7 days) to induce differentiation.
- Stimulated cells with interferon gamma and muramyl dipeptide to induce TNF production, followed by treatment with Platelet-activating factor (PAF) at concentrations ranging from 10(-16) to 10(-6) M.
- Monitored TNF production and intracellular calcium ([Ca2+]i) mobilization as indicators of cellular response.
Main Results:
- Undifferentiated HL-60 cells and those differentiated for 4 days showed no significant TNF response to PAF, despite baseline TNF production.
- By day 5 of differentiation, HL-60 cells exhibited a 2.5-fold increase in TNF production in response to PAF, with a single peak at 10(-8) M.
- Cells differentiated for 6-7 days displayed a bimodal TNF response to PAF, with peaks at 10(-14) M and 10(-7) M, mirroring responses seen in less dense monocyte subsets.
- Responsiveness to PAF in terms of [Ca2+]i mobilization was observed earliest at 4 days (10(-7) M) and became bimodal by day 7 (10(-14) and 10(-7) M).
Conclusions:
- Myeloid cell differentiation towards the monocyte lineage progressively enhances responsiveness to Platelet-activating factor (PAF).
- Acquired responsiveness includes increased intracellular calcium mobilization and augmented cytokine (TNF) production.
- The observed heterogeneity in normal monocyte responses to PAF may be attributed to their distinct differentiation or maturation stages.
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