title: Enhancement of Inducible Nitric Oxide Synthase Activity by Low Molecular Weight Peptides Derived from Protamine: A Potential Therapy for Chronic Rhinosinusitis.
abstract: Nitric oxide (NO) is a key immune defense agent that is produced from l-arginine in the airways by leukocytes and airway epithelial cells, primarily via inducible nitric oxide synthase (iNOS). Deficiencies in nasal NO levels have been associated with diseases such as primary ciliary dyskinesia and chronic rhinosinusitis. Herein, we demonstrate a proof-of-concept regarding a potential new therapeutic approach for such disorders. We show that arginine-rich low molecular weight peptides (LMWPs) derived from the FDA-approved protamine (obtained from salmon sperm) are effective at significantly raising NO production in both RAW 264.7 mouse macrophage and LA4 mouse epithelial cell lines. LMWP is produced using a stable, easily produced immobilized thermolysin gel column followed by size-exclusion purification. Monomeric l-arginine induces concentration-dependent increases in NO production in stimulated RAW 264.7 and LA4 cells, as measured by stable nitrite in the cell media. In stimulated RAW 264.7 cells, LMWP significantly increases iNOS expression and total NO production 12-24 h post-treatment compared to cells given equivalent levels of monomeric l-arginine. For stimulated LA4 cells, LMWPs are effective in significantly increasing NO production compared to equivalent l-arginine monomer concentrations over 24 h but do not substantially enhance iNOS expression. The use of the arginase inhibitor S-boronoethyl-l-cysteine in combination with LMWPs results in even higher NO production by stimulated RAW 264.7 cells and LA4 cells. Increases in NO due to LMWPs, compared to l-arginine, occur only after 4 h, which may be due to iNOS elevation rather than increased substrate availability.
authors.affiliation: †Department of Chemistry, University of Michigan, 930 North University, Ann Arbor, Michigan 48109, United States.
authors.forename: Anant S
authors.initials: AS
authors.lastname: Balijepalli
authors.affiliation: ‡Department of Pediatrics and Communicable Diseases, University of Michigan Health System, 1150 West Medical Center, Ann Arbor, Michigan 48109, United States.
authors.forename: Adam T
authors.initials: AT
authors.lastname: Comstock
authors.affiliation: †Department of Chemistry, University of Michigan, 930 North University, Ann Arbor, Michigan 48109, United States.
authors.forename: Xuewei
authors.initials: X
authors.lastname: Wang
authors.affiliation: †Department of Chemistry, University of Michigan, 930 North University, Ann Arbor, Michigan 48109, United States.
authors.forename: Gary C
authors.initials: GC
authors.lastname: Jensen
authors.affiliation: ‡Department of Pediatrics and Communicable Diseases, University of Michigan Health System, 1150 West Medical Center, Ann Arbor, Michigan 48109, United States.
authors.forename: Marc B
authors.initials: MB
authors.lastname: Hershenson
authors.affiliation: §Department of Otolaryngology, University of Michigan Health System, 1500 East Medical Center, Ann Arbor, Michigan 48109, United States.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Zacharek
authors.affiliation: ‡Department of Pediatrics and Communicable Diseases, University of Michigan Health System, 1150 West Medical Center, Ann Arbor, Michigan 48109, United States.
authors.forename: Umadevi S
authors.initials: US
authors.lastname: Sajjan
authors.affiliation: †Department of Chemistry, University of Michigan, 930 North University, Ann Arbor, Michigan 48109, United States.
authors.forename: Mark E
authors.initials: ME
authors.lastname: Meyerhoff
mesh_terms: D000818:Animals; D001119:Arginase / Q000378:metabolism; D001120:Arginine / Q000378:metabolism; D002460:Cell Line; D003545:Cysteine / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D010455:Peptides / Q000627:therapeutic use*; D011479:Protamines / Q000627:therapeutic use*; D065631:Rhinitis, Allergic / Q000188:drug therapy* / Q000378:metabolism
chemical_list: D009573:Nitrites; D010455:Peptides; D011479:Protamines; D009569:Nitric Oxide; D001120:Arginine; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D001119:Arginase; D003545:Cysteine
keywords: arginase; cell-penetrating peptides; chronic rhinosinusitis; cystic fibrosis; iNOS; nitric oxide; protamine; s-boronoethyl-l-cysteine