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Published on: January 7, 2020
Nafamostat mesilate suppresses NF-kappaB activation and NO overproduction in LPS-treated macrophages
Soichi Noguchi1, Mikiya Nakatsuka, Hideki Konishi
1Department of Obstetrics and Gynecology, Okayama University Medical School, 2-5-1 Shikata, Okayama Okayama 700-8558, Japan.
Abstract:
Nafamostat mesilate (NM), a clinically used serine protease inhibitor, suppressed the overproduction of nitric oxide (NO) and the expression of inducible nitric oxide synthase (iNOS) in RAW264.7 murine macrophages treated with lipopolysaccharide (LPS, 100 ng/ml); however, it had little effect on endothelial NOS (eNOS) in human umbilical vein endothelial cells (HUVEC). Electrophoretic mobility shift assay (EMSA) revealed that LPS activated nuclear factor-kappaB (NF-kappaB) in RAW264.7 cells and that this activation was suppressed by nafamostat mesilate. Western blotting showed that nafamostat mesilate suppressed the phosphorylation and degradation of inhibitor kappaB-alpha (IkappaB-alpha), which holds NF-kappaB in the cytoplasm in an inactivated state. Our observations suggest that nafamostat mesilate is a candidate agent for various diseases such as ischemia-reperfusion, graft rejection, inflammatory diseases, and autoimmune diseases, in which iNOS and/or NF-kappaB are upregulated.
Insights
Nafamostat mesilate effectively reduces nitric oxide (NO) overproduction and inducible nitric oxide synthase (iNOS) expression by inhibiting nuclear factor-kappaB (NF-kappaB) activation. This suggests its potential for treating inflammatory and autoimmune diseases.
Area of Science:
- Pharmacology
- Immunology
- Molecular Biology
Background:
- Nitric oxide (NO) overproduction and inducible nitric oxide synthase (iNOS) expression are implicated in various inflammatory and autoimmune diseases.
- Nuclear factor-kappaB (NF-kappaB) is a key transcription factor regulating inflammatory responses.
Purpose of the Study:
- To investigate the effect of nafamostat mesilate (NM) on nitric oxide (NO) production and NF-kappaB activation in inflammatory models.
- To explore the potential of NM as a therapeutic agent for diseases involving iNOS and NF-kappaB upregulation.
Main Methods:
- RAW264.7 murine macrophages and human umbilical vein endothelial cells (HUVEC) were used.
- Lipopolysaccharide (LPS) was used to induce inflammation.
- Nitric oxide (NO) production and iNOS expression were measured.
- Electrophoretic mobility shift assay (EMSA) was used to assess NF-kappaB activation.
- Western blotting was performed to analyze IkappaB-alpha phosphorylation and degradation.
Main Results:
- Nafamostat mesilate (NM) suppressed NO overproduction and iNOS expression in LPS-treated RAW264.7 cells.
- NM showed minimal effect on endothelial NOS (eNOS) in HUVEC.
- LPS-induced NF-kappaB activation in RAW264.7 cells was inhibited by NM.
- NM suppressed the phosphorylation and degradation of IkappaB-alpha, preventing NF-kappaB translocation.
Conclusions:
- Nafamostat mesilate (NM) inhibits iNOS expression and NF-kappaB activation, likely by stabilizing IkappaB-alpha.
- NM demonstrates potential as a therapeutic candidate for inflammatory diseases, ischemia-reperfusion injury, graft rejection, and autoimmune conditions characterized by iNOS and/or NF-kappaB upregulation.
