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Mitogen-activated protein kinase pathway mediates DBP-maf-induced apoptosis in RAW 264.7 macrophages
Kiranmai Gumireddy1, C Damodar Reddy, Narasimha Swamy
1Women and Infants' Hospital, Brown University, Providence, Rhode Island 02905, USA.
Abstract:
Vitamin D-binding protein-macrophage-activating factor (DBP-maf) is derived from serum vitamin D binding protein (DBP) by selective deglycosylation during inflammation. In the present study, we investigated the effect of DBP-maf on RAW 264.7 macrophages and the underlying intracellular signal transduction pathways. DBP-maf increased proapoptotic caspase-3, -8, and -9 activities and induced apoptosis in RAW 264.7 cells. However, DBP, the precursor to DBP-maf did not induce apoptosis in these cells. Cell cycle analysis of DBP-maf-treated RAW 264.7 cells revealed growth arrest with accumulation of cells in sub-G(0)/G(1) phase. We also investigated the role of mitogen-activated protein kinase (MAPK) pathways in the DBP-maf-induced apoptosis of RAW264.7 cells. DBP-maf increased the phosphorylation of p38 and JNK1/2, while it decreased the ERK1/2 phosphorylation. Treatment with the p38 MAPK inhibitor, SB202190, attenuated DBP-maf-induced apoptosis. PD98059, a MEK specific inhibitor, did not show a significant inhibition of apoptosis induced by DBP-maf. Taken together, these results suggest that the p38 MAPK pathway plays a crucial role in DBP-maf-mediated apoptosis of macrophages. Our studies indicate that, during inflammation DBP-maf may function positively by causing death of the macrophages when activated macrophages are no longer needed at the site of inflammation. In summary, we report for the first time that DBP-maf induces apoptosis in macrophages via p38 and JNK1/2 pathway.
Insights
Vitamin D-binding protein-macrophage-activating factor (DBP-maf) induces apoptosis in macrophages by activating the p38 and JNK pathways. This finding suggests DBP-maf plays a role in regulating immune responses during inflammation.
Area of Science:
- Immunology
- Cell Biology
- Molecular Biology
Background:
- Vitamin D-binding protein-macrophage-activating factor (DBP-maf) is a modified form of serum vitamin D binding protein (DBP) generated during inflammation.
- The precise role and mechanisms of DBP-maf in cellular processes, particularly macrophage apoptosis, remain incompletely understood.
Purpose of the Study:
- To investigate the effects of DBP-maf on RAW 264.7 macrophages.
- To elucidate the intracellular signaling pathways involved in DBP-maf-induced apoptosis.
Main Methods:
- Treatment of RAW 264.7 macrophages with DBP-maf and DBP.
- Assays for caspase activity, cell cycle analysis, and Western blotting to assess protein phosphorylation.
- Inhibition studies using p38 MAPK and MEK inhibitors (SB202190 and PD98059, respectively).
Main Results:
- DBP-maf, but not DBP, significantly increased proapoptotic caspase activity and induced apoptosis in macrophages.
- DBP-maf treatment led to cell cycle arrest at the sub-G(0)/G(1) phase.
- DBP-maf enhanced the phosphorylation of p38 and JNK1/2, while decreasing ERK1/2 phosphorylation.
- Inhibition of p38 MAPK attenuated DBP-maf-induced apoptosis, whereas MEK inhibition had no significant effect.
Conclusions:
- DBP-maf induces apoptosis in macrophages through the activation of the p38 and JNK mitogen-activated protein kinase (MAPK) pathways.
- The p38 MAPK pathway is crucial for DBP-maf-mediated macrophage apoptosis.
- DBP-maf may play a regulatory role in inflammation by eliminating activated macrophages when they are no longer required.