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Induction of apoptosis in B lineage cells by activin A derived from macrophages
T Nishihara1, Y Ohsaki, N Ueda
1Department of Oral Science, National Institute of Health, Tokyo, Japan.
Abstract:
A factor produced by P388D1 cell line murine macrophages showed a profound suppressive effect on the in vitro proliferation of B lineage cells. It was purified to homogeneity from conditioned media of P388D1 cells stimulated with phorbol 12-myristate 13-acetate for 48 h by a three-step procedure. The purified factor gave a single band of protein with a molecular mass of 16 kD on SDS-polyacrylamide gel electrophoresis. We show here that exposure of B lineage cells to this factor results in the induction of a cytotoxic effect and a significant increase in the proportion of fragmented DNA. DNA fragmentation was detected in B lineage cells after 3 h culture with the factor in the quantitative colorimetric determination. The mechanism of cell death was characterized by a ladder-like electrophoretic pattern of degraded chromosomal DNA, indicating that the factor induces apoptosis. The NH2-terminal amino acid sequence of this factor was identical with that of activin A over the 26 amino acid residues identified. We sought to determine whether apoptosis could be modulated by two kinds of inhibitor of protein kinases, H7 and HA1004, in concentrations that are below their toxicity limits. Apoptosis induced by the factor was suppressed by H7 but was relatively unaffected by HA1004. These findings suggest that the signals by protein kinases may regulate apoptotic B cell death by the factor activin A, derived from macrophages.
Insights
Macrophages produce a factor, identified as activin A, that induces apoptosis in B lineage cells. Protein kinase signaling pathways regulate this macrophage-derived factor-induced B cell death.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Macrophages play a crucial role in regulating immune responses.
- B cell proliferation and survival are critical for adaptive immunity.
- Understanding the mechanisms of B cell regulation is essential for immune system research.
Purpose of the Study:
- To investigate the effect of a macrophage-derived factor on B lineage cell proliferation.
- To elucidate the mechanism of cell death induced by this factor.
- To explore the role of protein kinases in mediating this process.
Main Methods:
- Purification of a suppressive factor from P388D1 macrophage cell line conditioned media.
- Analysis of the factor's effect on B lineage cell proliferation and DNA fragmentation.
- Characterization of cell death mechanism using DNA electrophoresis.
- NH2-terminal amino acid sequencing to identify the factor.
- Assessment of apoptosis modulation using protein kinase inhibitors (H7 and HA1004).
Main Results:
- A 16 kD protein factor, identified as activin A, was purified.
- This factor induced cytotoxicity and DNA fragmentation in B lineage cells, indicative of apoptosis.
- Apoptosis induced by activin A was suppressed by H7, but not HA1004.
- Protein kinase signaling pathways appear to regulate this apoptotic process.
Conclusions:
- Macrophage-derived activin A induces apoptosis in B lineage cells.
- Protein kinase signaling, particularly pathways sensitive to H7, plays a role in regulating this apoptosis.
- These findings shed light on the intricate mechanisms of immune cell regulation by macrophages.