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Doxorubicin (DOX) is a potent available antitumor agent; however, its clinical use is limited because of its cardiotoxicity. Cell death is a key component in DOX-induced cardiotoxicity, but its mechanisms are elusive. Here, we explore the role of superoxide, nitric oxide (NO), and peroxynitrite in DOX-induced cell death using both in vivo and in vitro models of cardiotoxicity. Western blot analysis, real-time PCR, immunohistochemistry, flow cytometry, fluorescent microscopy, and biochemical assays were used to determine the markers of apoptosis/necrosis and sources of NO and superoxide and their production. Left ventricular function was measured by a pressure-volume system. We demonstrated increases in myocardial apoptosis (caspase-3 cleavage/activity, cytochrome c release, and TUNEL), inducible NO synthase (iNOS) expression, mitochondrial superoxide generation, 3-nitrotyrosine (NT) formation, matrix metallo-proteinase (MMP)-2/MMP-9 gene expression, poly(ADP-ribose) polymerase activation [without major changes in NAD(P)H oxidase isoform 1, NAD(P)H oxidase isoform 2, [p22.sup.phox], [p40.sup.phox], [p47.sup.phox], [p67.sup.phox], xanthine oxidase, endothelial NOS, and neuronal NOS expression] and decreases in myocardial contractility, catalase, and glutathione peroxidase activities 5 days after DOX treatment to mice. All these effects of DOX were markedly attenuated by peroxynitrite scavengers. Doxorubicin dose dependently increased mitochondrial superoxide and NT generation and apoptosis/necrosis in cardiac-derived H9c2 cells. DOX- or peroxynitrite-induced apoptosis/necrosis positively correlated with intracellular NT formation and could be abolished by peroxynitrite scavengers. DOX-induced cell death and NT formation were also attenuated by selective iNOS inhibitors or in iNOS knockout mice. Various NO donors when coadministered with DOX but not alone dramatically enhanced DOX-induced cell death with concomitant increased NT formation. DOX-induced cell death was also attenuated by cell-permeable SOD but not by cell-permeable catalase, the xanthine oxidase inhibitor allopurinol, or the NADPH oxidase inhibitors apocynine or diphenylene iodonium. Thus, peroxynitrite is a major trigger of DOX-induced cell death both in vivo and in vivo, and the modulation of the pathways leading to its generation or its effective neutralization can be of significant therapeutic benefit. heart failure; apoptosis; inducible nitric oxide synthase; hemodynamics

Reversible cerebral vasoconstriction syndrome (RCVS) is characterized by recurrent severe thunderclap headache and segmental vasoconstriction of cerebral arteries that resolves by 3 months. RCVS can either occur spontaneously or relate to an exogenous trigger, but the precise pathophysiology of RCVS remains unknown. Postulated mechanisms include small vessel and endothelial dysfunction, mitochondrial dysfunction with oxidative stress, hormonal and biochemical factors, and genetic predisposition. A prior history of migraine has been reported in 17%–40% of cases, suggesting that migraine and RCVS share mechanisms. Most RCVS patients have a favorable prognosis; however, for some patients, the condition may result in permanent disability or even death in a small minority of patients, secondary to complications such as ischemic stroke or intracranial hemorrhage. There is no proven treatment, so the cornerstone of RCVS management remains largely supportive, with prescription of bed rest, analgesics, and the removal of precipitating factors. The majority of clinical series studying RCVS treatment have investigated the efficacy of calcium channel antagonists. Although prescription of triptans for migraine is not suitable for patients with a history of RCVS, lasmiditan, a highly selective 5‐HT1F receptor agonist, may be suitable in such cases. Typical segmental vasoconstriction of major vessels is not seen in approximately 30% of patients eventually diagnosed with RCVS; if this diagnosis is suspected, repeat imaging should be performed. Early diagnosis of RCVS is important, as it allows progression to be prevented by the prompt removal of precipitating factors, unnecessary diagnostic procedures to be avoided, and appropriate management to be commenced. [ABSTRACT FROM AUTHOR]

The heroin analogue 1-methyl-4-phenylpyridinium, [MPP.sup.+], both in vitro and in vivo, produces death of dopaminergic substantia nigral cells by inhibiting the mitochondrial NADH dehydrogenase multienzyme complex, producing a syndrome indistinguishable from Parkinson's disease. Similarly, a fragment of amyloid protein, A[[Beta].sub.1-42], is lethal to hippocampal cells, producing recent memory deficits characteristic of Alzheimer's disease. Here we show that addition of 4 mM D-[Beta]-hydroxybutyrate protected cultured mesencephalic neurons from [MPP.sup.+] toxicity and hippocampal neurons from A[[Beta].sub.1-42] toxicity. Our previous work in heart showed that ketone bodies, normal metabolites, can correct defects in mitochondrial energy generation. The ability of ketone bodies to protect neurons in culture suggests that defects in mitochondrial energy generation contribute to the pathophysiology of both brain diseases. These findings further suggest that ketone bodies may play a therapeutic role in these most common forms of human neurodegeneration.

Background Providing ketone bodies to the brain can bypass metabolic blocks to glucose utilization and improve function in energy-starved neurons. For this, plasma ketones must be elevated well above the ≤0.2 mM default concentrations normally prevalent. Limitations of dietary methods currently used to produce therapeutic hyperketonemia have stimulated the search for better approaches. Method Described herein is a new way to produce therapeutic hyperketonemia, entailing prolonged oral administration of a potent ketogenic agent—ketone monoester (KME)—to a patient with Alzheimer's disease dementia and a pretreatment Mini-Mental State Examination score of 12. Results The patient improved markedly in mood, affect, self-care, and cognitive and daily activity performance. The KME was well tolerated throughout the 20-month treatment period. Cognitive performance tracked plasma β-hydroxybutyrate concentrations, with noticeable improvements in conversation and interaction at the higher levels, compared with predose levels. Conclusion KME-induced hyperketonemia is robust, convenient, and safe, and the ester can be taken as an oral supplement without changing the habitual diet. [ABSTRACT FROM AUTHOR]

Rapid inactivation of metabolism is essential for accurately determining the concentrations of metabolic intermediates in the in vivo state. We compared a broad spectrum of energetic intermediate metabolites and neurotransmitters in brains obtained by microwave irradiation to those obtained by freeze blowing, the most rapid method of extracting and freezing rat brain. The concentrations of many intermediates, cytosolic free NAD( P)+/ NAD( P) H ratios, as well as neurotransmitters were not affected by the microwave procedure. However, the brain concentrations of ATP were about 30% lower, whereas those of ADP, AMP, and GDP were higher in the microwave-irradiated compared with the freeze-blown brains. In addition, the hydrolysis of approximately 1 μmol/g of ATP, a major in vivo Mg2+-binding site, was related to approximately five-fold increase in free [Mg2+] (0.53 ± 0.07 mM in freeze blown vs. 2.91 mM ± 0.48 mM in microwaved brains), as determined from the ratio [citrate]/[isocitrate]. Consequently, many intracellular properties, such as the phosphorylation potential and the ∆G' of ATP hydrolysis were significantly altered in microwaved tissue. The determinations of some glycolytic and TCA cycle metabolites, the phosphorylation potential, and the ∆G' of ATP hydrolysis do not represent the in vivo state when using microwave-fixed brain tissue. [ABSTRACT FROM AUTHOR]

Ketosis, meaning elevation of D-β-hydroxybutyrate (R-3-hydroxybutyrate) and acetoacetate, has been central to starving man's survival by providing nonglucose substrate to his evolutionarily hypertrophied brain, sparing muscle from destruction for glucose synthesis. Surprisingly, D-β-hydroxybutyrate (abbreviated "βOHB") may also provide a more efficient source of energy for brain per unit oxygen, supported by the same phenomenon noted in the isolated working perfused rat heart and in sperm. It has also been shown to decrease cell death in two human neuronal cultures, one a model of Alzheimer's and the other of Parkinson's disease. These observations raise the possibility that a number of neurologic disorders, genetic and acquired, might benefit by ketosis. Other beneficial effects from βOHB include an increased energy of ATP hydrolysis (ΔG') and its linked ionic gradients. This may be significant in drug-resistant epilepsy and in injury and anoxic states. The ability of βOHB to oxidize co-enzyme Q and reduce NADP[sup +] may also be important in decreasing free radical damage. Clinical maneuvers for increasing blood levels of βOHB to 2-5 mmol may require synthetic esters or polymers of βOHB taken orally, probably 100 to 150 g or more daily. This necessitates advances in food-science technology to provide at least enough orally acceptable synthetic material for animal and possibly subsequent clinical testing. The other major need is to bring the technology for the analysis of multiple metabolic "phenotypes" up to the level of sophistication of the instrumentation used, for example, in gene science or in structural biology. This technical strategy will be critical to the characterization of polygenic disorders by enhancing the knowledge gained from gene analysis and from the subsequent steps and modifications of the protein products themselves. [ABSTRACT FROM AUTHOR]