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COM crystals activate the p38 mitogen-activated protein kinase signal transduction pathway in renal epithelial cells
Hari K Koul1, Mani Menon, Lakshmi S Chaturvedi
1Biochemistry and Molecular Biology Laboratory, Vattikuti Urology Institute, Henry Ford Health Sciences Center, Detroit, Michigan 48202, USA. hkoul1@hfhs.org
Abstract:
Interaction of calcium oxalate monohydrate (COM) crystals with renal cells has been shown to result in altered gene expression, DNA synthesis, and cell death. In the current study the role of a stress-specific p38 MAP kinase-signaling pathway in mediating these effects of COM crystals was investigated. Exposure of cells to COM crystals (20 microg/cm(2)) rapidly stimulated strong phosphorylation and activation of p38 mitogen-activated protein kinase (p38 MAP kinase) and re-initiation of DNA synthesis. Inhibition of COM crystal binding to the cells by heparin blocked the effects of COM crystals on p38 MAPK activation. We also show that specific inhibition of p38 MAPK by 4-(4-fluorophenyl)-2-(4-methylsulfonylphenyl)-5-(4-pyridyl) imidazole (SB203580) or by overexpression of a dominant negative mutant of p38 MAP kinase abolishes COM crystal-induced re-initiation of DNA synthesis. The inhibition is dose-dependent and correlates with in situ activity of native p38 MAP kinase, determined as mitogen-activated protein kinase-activated protein kinase-2 (MAPKAP kinase-2) activity in cell extracts. In summary, inhibiting activation of p38 MAPK pathway abrogated the DNA synthesis in response to COM crystals. These data are the first demonstrations of activation of the p38 MAPK signaling pathway by COM crystals and suggest that, in response to COM crystals, this pathway transduces critical signals governing the re-initiation of DNA synthesis in renal epithelial cells.
Insights
Calcium oxalate monohydrate (COM) crystals activate the p38 MAP kinase pathway in kidney cells, triggering DNA synthesis. Inhibiting this pathway blocks COM-induced DNA re-initiation, revealing its critical role in kidney stone formation.
Area of Science:
- Nephrology
- Molecular Biology
- Cell Biology
Background:
- Calcium oxalate monohydrate (COM) crystal interaction with renal cells alters gene expression, DNA synthesis, and cell death.
- The p38 MAP kinase (MAPK) pathway is a stress-specific signaling cascade implicated in cellular responses to injury.
Purpose of the Study:
- To investigate the role of the p38 MAP kinase signaling pathway in mediating the effects of COM crystals on renal cells.
- To determine if p38 MAPK activation is essential for COM crystal-induced DNA synthesis.
Main Methods:
- Exposure of renal cells to COM crystals and assessment of p38 MAPK phosphorylation and activation.
- Inhibition of COM crystal binding using heparin.
- Pharmacological inhibition of p38 MAPK using SB203580 and genetic inhibition via dominant-negative p38 MAPK mutant.
- Measurement of mitogen-activated protein kinase-activated protein kinase-2 (MAPKAP kinase-2) activity as an indicator of p38 MAPK activity.
Main Results:
- COM crystals rapidly stimulated p38 MAPK phosphorylation and activation, leading to DNA synthesis re-initiation.
- Heparin blockade of COM crystal binding inhibited p38 MAPK activation.
- Specific inhibition of p38 MAPK abolished COM crystal-induced DNA synthesis in a dose-dependent manner.
- Inhibition of p38 MAPK activity correlated with reduced DNA synthesis.
Conclusions:
- The p38 MAP kinase signaling pathway is activated by COM crystals in renal epithelial cells.
- p38 MAPK activation is critical for the re-initiation of DNA synthesis in response to COM crystals.
- This pathway transduces signals that govern renal cell DNA synthesis following COM crystal interaction, suggesting a potential therapeutic target for kidney stones.