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Certain S-substituted isothioureas not only inhibit NO synthase catalytic activity but also decrease translation and
L H Wei1, N Arabolos, L J Ignarro
1Department of Molecular and Medical Pharmacology, UCLA School of Medicine, Los Angeles, California 90095-1735, USA.
Nitric Oxide : Biology and Chemistry
|September 10, 1998
Summary
S-Ethylisothiourea (EITU) is a potent inhibitor of inducible nitric oxide synthase (iNOS) in cell culture. Unlike AEITU, EITU inhibits NO production without affecting iNOS induction, making it suitable for research.
Area of Science:
- Biochemistry
- Pharmacology
- Cell Biology
Background:
- Inducible nitric oxide synthase (iNOS) plays a crucial role in cellular processes.
- Developing specific inhibitors for iNOS is essential for research applications.
- Existing inhibitors like NG-methylarginine and aminoguanidine have limitations.
Purpose of the Study:
- To identify potent and selective inhibitors of iNOS for cell culture.
- To evaluate the efficacy and mechanism of S-substituted isothioureas as iNOS inhibitors.
- To differentiate between direct catalytic inhibition and interference with iNOS induction.
Main Methods:
- Cultured RAW 264.7 macrophages stimulated with lipopolysaccharide (LPS).
- Assay of nitric oxide (NO) production via 24-h accumulation in cell culture media.
- Analysis of iNOS induction using Northern blot for mRNA, pulse labeling for translation, and pulse-chase for protein degradation.
Main Results:
- S-Ethylisothiourea (EITU) and S-aminoethylisothiourea (AEITU) showed potent inhibition of NO production (EC50: 10 µM and 30 µM, respectively).
- EITU inhibited NO production without affecting iNOS mRNA transcription, protein translation, or degradation.
- AEITU interfered with iNOS induction by inhibiting translation and accelerating protein degradation.
Conclusions:
- EITU is a selective inhibitor of iNOS catalytic activity, suitable for cell culture studies.
- AEITU's interference with iNOS induction mechanisms limits its use for solely assessing catalytic inhibition.
- These findings provide valuable tools for researchers studying NO signaling pathways.