Reactive oxygen species (ROS) and lipid peroxidation (LPO) have been associated with numerous diseases that few other pathological factors can match, including aging, neoplasia, trauma, and ischemia-reperfusion injury (Halliwell and Gutteridge 1999). The mechanism of involvement of LPO has been an area of intense research aiming to prevent, slow down, and even reverse the development of various diseases. In the case of spinal cord injury, it is well established that LPO plays an important role in neuronal degeneration, cell death, and overall functional deficits (Hall 1989(Hall , 1991Hall and Braughler 1993). This is believed due in part to that fact that neuronal cells contain a relatively large proportion of polyunsaturated fatty acids and are rich in mitochondria, both of which are a potential target and source of free radicals. Because of these unique features, the CNS is particularly vulnerable to oxidative injury. In spite of strong evidence suggesting that post-trauma oxidative stress plays a critical role in the pathogenesis of spinal cord injury, conventional strategies aiming to scavenge free radicals have largely failed to produce any effective treatment that can curtail oxidative injury. Hence, further understanding of the mechanisms of oxidative stress and identification of a novel and more effective target is highly warranted and desirable.In addition to the much studied ROS, highly reactive a,b-unsaturated aldehydes, including malondialdehyde, 4-hydroxynonenal (HNE), and acrolein, are produced as a byproduct of LPO (Witz 1989;Esterbauer et al. 1991;Uchida 1999;O'Brien et al. 2005). Among them, acrolein has been shown to be by far the most reactive with various biomolecules including proteins, DNA, and glutathione, and reacts 110-150 times faster with glutathione than HNE or Abbreviations used: AD, Alzheimer's disease; CAP, compound action potential conduction; DTNB, 5,5¢-Dithiobis (2-nitrobenzoic acid); HE, dihydroethidium; HNE, 4-hydroxynonenal; LDH, lactate dehydrogenase; LPO, lipid peroxidation; PB, phosphate buffer; PBS, phosphatebuffered saline; ROS, reactive oxygen species; SOD, superoxide dismutase; TMR, tetramethyl rhodamine dextran. AbstractWe have previously shown that acrolein, a lipid peroxidation byproduct, is significantly increased following spinal cord injury in vivo, and that exposure to neuronal cells results in oxidative stress, mitochondrial dysfunction, increased membrane permeability, impaired axonal conductivity, and eventually cell death. Acrolein thus may be a key player in the pathogenesis of spinal cord injury, where lipid peroxidation is known to be involved. The current study demonstrates that the acrolein scavenger hydralazine protects against not only acrolein-mediated injury, but also compression in guinea pig spinal cord ex vivo. Specifically, hydralazine (500 lmol/L to 1 mmol/L) can significantly alleviate acrolein (100-500 lmol/ L)-induced superoxide production, glutathione depletion, mitochondrial dysfunction, loss of membrane integrity, and reduced compound act...

The pathophysiology of spinal cord injury (SCI) is characterized by the initial, primary injury followed by secondary injury processes in which oxidative stress is a critical component. Secondary injury processes not only exacerbate pathology at the site of primary injury, but also result in spreading of injuries to the adjacent, otherwise healthy tissue. The lipid peroxidation byproduct acrolein has been implicated as one potential mediator of secondary injury. In order to further and rigorously elucidate the role of acrolein in secondary injury, a unique ex vivo model is utilized to isolate the detrimental effects of mechanical injury from toxins such as acrolein that are produced endogenously following SCI. We demonstrate that: 1) acrolein-lys adducts are capable of diffusing from compressed tissue to adjacent, otherwise uninjured tissue; 2) secondary injury by itself produces significant membrane damage and increased superoxide production; and 3) these injuries are significantly attenuated by the acrolein scavenger hydralazine. Furthermore, hydralazine treatment results in significantly less membrane damage 2 hours following compression injury, but not immediately after. These findings support our hypothesis that, following SCI, acrolein is increased to pathologic concentrations, contributes significantly to secondary injury, and thus represents a novel target for scavenging to promote improved recovery.

It has long been established that oxidative stress plays a critical role in the pathophysiology of spinal cord injury, and represents an important target of therapeutic intervention following the initial trauma. However, free radical scavengers have been largely ineffective in clinical trials, and as such a novel target to attenuate oxidative stress is highly warranted. In addition to free radicals, peroxidation of lipid membranes following spinal cord injury (SCI) produces reactive aldehydes such as acrolein. Acrolein is capable of depleting endogenous antioxidants such as glutathione, generating free radicals, promoting oxidative stress, and damaging proteins and DNA. Acrolein has a significantly longer half-life than the transient free radicals, and thus may represent a potentially better target of therapeutic intervention to attenuate oxidative stress. There is growing evidence, from our lab and others, to suggest that reactive aldehydes such as acrolein play a critical role in oxidative stress and SCI. The focus of this review is to summarize the cellular and biochemical mechanisms of acrolein-induced membrane damage, mitochondrial injury, oxidative stress, cell death, and functional loss. Evidence will also be presented to suggest that acrolein scavenging may be a novel means of therapeutic intervention to attenuate oxidative stress and improve recovery following traumatic SCI.

The integrity of the neuronal membrane is critical for its function as well as survival, and ineffective repair of damaged membranes may be one of the key factors underlying the neuronal degeneration and overall functional loss that occurs after spinal cord injury and traumatic brain injury. Previously, we showed that polyethylene glycol (PEG) can reseal axonal membranes following compression in isolated guinea pig spinal cord white matter. We now report that 10 mM PEG can also significantly enhance membrane resealing following transection in the clinically relevant conditions of low extracellular Ca(2+) and low temperature. Such beneficial effects were demonstrated both functionally, through membrane potential measured by double sucrose gap apparatus, and anatomically, through horseradish peroxidase and tetramethyl rhodamine dextran dye exclusion assays. We further noted that axons with small diameters preferentially benefited from PEG-mediated axolemmal resealing. Using atomic force microscopy, we further showed that PEG can effectively reduce neuronal membrane surface tension. We hypothesize that PEG may promote axolemmal resealing by increasing membrane line tension and reducing membrane tension, thus creating conditions more favorable to membrane resealing. In summary, these studies suggest that PEG is effective under the clinically relevant conditions of low Ca(2+) and temperature, and thus has the potential to be used in combination with other more established interventions in spinal cord and traumatic brain injury.

Objective: Polyethylene glycol (PEG), a hydrophilic polymer, can immediately repair neuronal membranes and inhibit free radical production following trauma. The aim of this study is to examine whether PEG can directly repair mitochondria in the event of trauma. Method: Purified brain mitochondria from guinea pigs were used. Mitochondrial function was assessed by biochemical methods and structural changes were observed by both fluorescence light microscopy and coherent anti-Stokes Raman scattering microscopy. Results: We present evidence suggesting that PEG is capable of directly reducing injury to mitochondria independent of plasma membrane repair. Specifically, the suppression of oxygen consumption rate of purified mitochondria due to H₂O₂ and/or calcium can be significantly reversed by 12.5 mM PEG. PEG also significantly reduced mitochondrial swelling due to similar injury. Furthermore, we have shown that such PEG-mediated mitochondrial protection is dependent on the molecular weight of PEG, suggesting a direct physical blockade of mitochondrial permeability transitional pore by PEG. Conclusion: These findings, coupled with previous evidence that PEG enters the cytosol following mechanical trauma, strongly indicate that there are at least 2 avenues of PEG-mediated cytoprotection in mechanically injured spinal cords: repair of plasma membrane and protection of mitochondria.