title: Acute arterial thrombosis causes endothelial dysfunction: a new paradigm for thrombolytic therapy.
abstract: OBJECTIVE
  The goals of this study were to delineate the time course of endothelial dysfunction after arterial thrombosis, to determine the cause of endothelial dysfunction in this setting, and to determine whether modulating standard thrombolytic therapy would ameliorate the thrombosis-mediated endothelial dysfunction.
  
  
  METHODS
  Male adult rats underwent infrarenal aortic occlusion by means of clip ligature to induce arterial thrombosis. After 30 minutes, 1, 2, and 3 hours, ring segments from the infrarenal aorta were harvested and placed into physiologic buffer baths. With the use of a force transducer, both endothelial-dependent relaxation (EDR) and endothelial-independent relaxation (EIR) were measured. Endothelial function and presence were determined by means of factor VIII immunohistochemical staining. Endothelial morphology was evaluated with scanning electron microscopy (SEM). Nitric oxide (NO) levels were determined with a chemiluminescent assay of its nitrite/nitrate metabolites (NO(x)). Standard thrombolytic therapy with urokinase (UK) was infused into thrombosed aortic ring segments and compared with UK supplemented with both low-dose L -arginine (2 mmol) and high-dose L -arginine (20 mmol).
  
  
  RESULTS
  Arterial thrombosis decreases EDR. The nadir of EDR occurs 1 hour after thrombosis (mean +/- SE, 13% +/- 6.4% vs 94% +/- 2.6% for controls, P <.005), with persistent lowering of EDR as long as 3 hours after thrombosis. EIR is preserved, and vasoconstriction with norepinephrine or potassium buffer is unaltered. Both endothelial function and presence (n = 6 per group) were documented by means of factor VIII immunohistochemistry. An intact monolayer of endothelium at all time intervals after thrombosis was revealed by means of SEM analysis. No differences between control and thrombosed specimens were revealed by means of the grading of SEM images. Local NO(x) levels were lower after 1 hour of thrombosis, with an increase higher than baseline values at 3 hours. The addition of low-dose L -arginine resulted in a minor increase in EDR. However, high-dose L -arginine resulted in a significant increase in EDR versus controls receiving UK alone (64% +/- 6.3% vs 38% +/- 4.4%, P <.05). Correspondingly, local NO(x) levels were 20-fold higher after the high-dose L -arginine supplementation when compared with UK thrombolysis alone (2.8 +/- 0.52 micromol/L vs 0.133 +/- 0.02 micromol/L, n = 6 samples/group, P <.005).
  
  
  CONCLUSIONS
  Acute arterial thrombosis causes endothelial dysfunction, without causing endothelial cell loss. Endothelial function reaches a nadir after 1 hour of thrombosis. EIR and vasoconstriction remain unaffected, indicating normal smooth muscle cell function. NO(x) levels suggest that NO levels are decreased acutely after thrombosis. Supplementing standard thrombolytic therapy with the NO precursor, l-arginine, ameliorates the endothelial dysfunction seen after acute thrombosis by increasing local NO production.
authors.affiliation: Vascular Surgery, Wilford Hall Medical Center, Lackland AFB, Tex, USA.
authors.forename: V S
authors.initials: VS
authors.lastname: Kashyap
authors.forename: T D
authors.initials: TD
authors.lastname: Reil
authors.forename: W S
authors.initials: WS
authors.lastname: Moore
authors.forename: T X
authors.initials: TX
authors.lastname: Hoang
authors.forename: H A
authors.initials: HA
authors.lastname: Gelabert
authors.forename: R E
authors.initials: RE
authors.lastname: Byrns
authors.forename: L J
authors.initials: LJ
authors.lastname: Ignarro
authors.forename: J A
authors.initials: JA
authors.lastname: Freischlag
mesh_terms: D000208:Acute Disease; D000818:Animals; D001157:Arterial Occlusive Diseases / Q000188:drug therapy*; D008297:Male; D009126:Muscle Relaxation; D009131:Muscle, Smooth, Vascular / Q000503:physiopathology; D009569:Nitric Oxide / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015912:Thrombolytic Therapy; D013927:Thrombosis / Q000188:drug therapy*
chemical_list: D009569:Nitric Oxide