Abstract
Nitroglycerin (NTG) is an important cardiovascular agent, but tolerance during continuous administration limits its clinical utility. Increased vascular superoxide production may mediate nitrate tolerance via a reduction in nitric oxide availability. Because superoxide anion and nitric oxide react avidly to form peroxynitrite, an aggressive cellular toxicant that nitrates protein tyrosine residues, we tested the hypotheses that protein nitration, indicative of peroxynitrite formation, occurs during vascular tolerance, and that protein nitration participates in tolerance development. Preincubation of rat thoracic aorta segments with NTG (22 μM, EC95 for 30 min) caused a significant shift in NTG relaxation response (EC50; 6.7 ± 1.7 versus 0.50 ± 0.13 μM, NTG versus vehicle,p < .05). After functional evaluations, tissues were fixed in formalin for immunohistochemistry and digital image analysis. NTG-induced vascular tolerance was associated with increased immunoprevalence of 3-nitrotyrosine (3NT, stable biomarker of protein nitration; 11.41 ± 2.48 versus 0.04 ± 0.02% positive pixels, NTG versus vehicle, p < .05). Staining was observed throughout vascular smooth muscle layers. Addition of 500 μM free tyrosine to the preincubation medium did not alter tolerance development (NTG EC50 6.5 ± 3.0 μM) but abolished 3NT immunoprevalence (0.16 ± 0.10%). No significant relationship between NTG potency and 3NT immunoprevalence was observed. These data support the hypothesis that protein nitration occurs during nitrate vascular tolerance, however, it apparently does not mediate this phenomenon.
Footnotes
-
Send reprint requests to: Dr. John Anthony Bauer, 412 Riffe Bldg., 500 West 12th Ave., Columbus, OH 43210-1291. E-mail:bauer.140{at}osu.edu
-
↵1 Supported in part by grants from the American Heart Association (Ohio-West Virginia Affiliate) and National Institutes of Health Grant HL059791.
- Abbreviations:
- NTG
- nitroglycerin
- NO
- nitric oxide
- O⨪2
- superoxide anion
- ONOO−
- peroxynitrite
- 3NT
- 3-nitro-l-tyrosine
- TYR
- tyrosine
- TNM
- tetranitromethane
- Received March 18, 1999.
- Accepted June 18, 1999.
- The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|