Isoketals and levuglandins are highly reactive γ-ketoaldehydes formed by oxygenation of arachidonic acid in settings of oxidative injury and cyclooxygenase activation, respectively. These compounds rapidly adduct to proteins via lysyl residues, which can alter protein structure/function. We examined whether pyridoxamine, which has been shown to scavenge α-ketoaldehydes formed by carbohydrate or lipid peroxidation, could also effectively protect proteins from the more reactive γ-ketoaldehydes. Pyridoxamine prevented adduction of ovalbumin and also prevented inhibition of RNase A and glutathione reductase activity by the synthetic γ-ketoaldehyde, 15-E 2 -isoketal. We identified the major products of the reaction of pyridoxamine with the 15-E 2 -isoketal, including a stable lactam adduct. Two lipophilic analogs of pyridoxamine, salicylamine and 5'O-pentylpyridoxamine, also formed lactam adducts when reacted with 15-E 2 -isoketal. When we oxidized arachidonic acid in the presence of pyridoxamine or its analogs, pyridoxamine-isoketal adducts were found in significantly greater abundance than the pyridoxamine-N-acyl adducts formed by α-ketoaldehyde scavenging. Therefore, pyridoxamine and its analogs appear to preferentially scavenge γ-ketoaldehydes. Both pyridoxamine and its lipophilic analogs inhibited the formation of lysyl-levuglandin adducts in platelets activated ex vivo with arachidonic acid. The two lipophilic pyridoxamine analogs provided significant protection against H 2 O 2 -mediated cytotoxicity in HepG2 cells. These results demonstrate the utility of pyridoxamine and lipophilic pyridoxamine analogs to assess the potential contributions of isoketals and levuglandins in oxidant injury and inflammation and suggest their potential utility as pharmaceutical agents in these conditions.Highly reactive γ-ketoaldehydes are formed via the cyclooxygenase pathway and by radicalcatalyzed lipid peroxidation. Prostaglandin H 2 , the product of the cyclooxygenase enzyme, rearranges in aqueous solution to form a number of eicosanoids, approximately 20% of which are the γ-ketoaldehydes levuglandin E 2 and D 2 . Lipid peroxidation yields a series of † This work was supported by National Institutes of Health Grants GM42056 (MERIT Award to L.J.R.), AG26119, GM15431 (to J . NIH Public Access Author ManuscriptBiochemistry. Author manuscript; available in PMC 2008 December 8. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript prostaglandin H 2 isomers that also rearrange to corresponding γ-ketoaldehydes, designated as isoketals (IsoK). These γ-ketoaldehydes (γKAs) react extremely rapidly with the lysyl residues of protein to form stable adducts, including a lysyl-lactam adduct and intermolecular crosslinks (1-4). Levels of γKA adducts significantly increase in pathological conditions including atherosclerosis, end-stage renal disease, and Alzheimer's Disease (5,6). Increased γKA adduct formation has also been characterized in experimental models of oxidative injury and inflammation, including carb...

A sixty-one-year old woman with a history of depression and idiopathic dystonia presented to the emergency department (ED) with fever, nausea, muscle rigidity, and redness in her extremities. The patient had seen various specialists for her dystonia during the preceding year and had tried multiple medication regimens, including benztropine and tizanidine. The patient described her home medications as tizanidine 2 mg three times daily, diazepam 2.5 mg daily, escitalopram 100 mg daily, rosuvastatin 10 mg daily and bupropion 300 mg nightly.At ED presentation her vital signs were: temperature 37.5°C (99.5F), blood pressure 157/76 mmHg, heart rate 105 beats per minute, respirations 24 per minute, and oxygen saturation of 100% on room air. Her physical examination was significant for diaphoresis, tachycardia, diffuse flushing, tremors in her upper and lower extremities, muscle rigidity, and hyperreflexia in the upper and lower extremities. The tremors were low amplitude and worse with intention, while the rigidity had cogwheeling features and was more notable in her lower extremities. Initial laboratory evaluation was significant for a creatine phosphokinase (CPK) of 1146 IU/L. Her electrocardiogram showed sinus tachycardia with normal QRS and QT intervals, and no evidence of acute ischemia.Two months earlier, the patient's neurologist had started her on amantadine 100 mg twice daily for dystonia. Three days prior to ED presentation, the patient abruptly stopped taking her amantadine on the advice of a pharmacist, secondary to bilateral lowerextremity edema. The following day, the patient presented to her primary care physician complaining of nausea, tremors, and diffuse flushing. The physician treated her for an allergic reaction with oral steroids and diphenhydramine. The patient's symptoms worsened over the next day with increasing tremors and rigidity, low-grade fever, nausea, and flushing, which prompted her presentation to the ED.