Efficacy of N-Acetyl Cysteine in Traumatic Brain Injury

In this study, using two different injury models in two different species, we found that early post-injury treatment with N-Acetyl Cysteine (NAC) reversed the behavioral deficits associated with the TBI. These data suggest generalization of a protocol similar to our recent clinical trial with NAC in blast-induced mTBI in a battlefield setting [1], to mild concussion from blunt trauma. This study used both weight drop in mice and fluid percussion injury in rats. These were chosen to simulate either mild or moderate traumatic brain injury (TBI). For mice, we used novel object recognition and the Y maze. For rats, we used the Morris water maze. NAC was administered beginning 30–60 minutes after injury. Behavioral deficits due to injury in both species were significantly reversed by NAC treatment. We thus conclude NAC produces significant behavioral recovery after injury. Future preclinical studies are needed to define the mechanism of action, perhaps leading to more effective therapies in man.     

N-Acetyl Cysteine Mitigates the Acute Effects of Cocaine-Induced Toxicity in Astroglia-Like Cells

Cocaine has a short half-life of only about an hour but its effects, predominantly on the central nervous system (CNS), are fairly long-lasting. Of all cells within the CNS, astrocytes may be the first to display cocaine toxicity owing to their relative abundance in the brain. Cocaine entry could trigger several early response changes that adversely affect their survival, and inhibiting these changes could conversely increase their rate of survival. In order to identify these changes and the minimal concentrations of cocaine that can elicit them in vitro, rat C6 astroglia-like cells were treated with cocaine (2–4 mM for 1h) and assayed for alterations in gross cell morphology, cytoplasmic vacuolation, viability, reactive oxygen species (ROS) generation, glutathione (GSH) levels, cell membrane integrity, F-actin cytoskeleton, and histone methylation. We report here that all of the above identified features are significantly altered by cocaine, and may collectively represent the key pathology underlying acute toxicity-mediated death of astroglia-like cells. Pretreatment of the cells with the clinically available antioxidant N-acetyl cysteine (NAC, 5 mM for 30 min) inhibited these changes during subsequent application of cocaine and mitigated cocaine-induced toxicity. Despite repeated cocaine exposure, NAC pretreated cells remained highly viable and post NAC treatment also increased viability of cocaine treated cells to a smaller yet significant level. We show further that this alleviation by NAC is mediated through an increase in GSH levels in the cells. These findings, coupled with the fact that astrocytes maintain neuronal integrity, suggest that compounds which target and mitigate these early toxic changes in astrocytes could have a potentially broad therapeutic role in cocaine-induced CNS damage.     

Determination of DNA Damage in Experimental Liver Intoxication and Role of N-Acetyl Cysteine

The present study aimed at detecting DNA damage and fragmentation as well as histone acetylation depending on oxidative stress caused by CCl₄ intoxication. Also, the protective role of N-acetyl cysteine, a precursor for GSH, in DNA damage is investigated. Sixty rats were used in this study. In order to induce liver toxicity, CCl₄ in was dissolved in olive oil (1/1) and injected intraperitoneally as a single dose (2 ml/kg). N-acetyl cysteine application (intraperitoneal, 50 mg/kg/day) was started 3 days prior to CCl₄ injection and continued during the experimental period. Control groups were given olive oil and N-acetyl cysteine. After 6 and 72 h of CCl₄ injection, blood and liver tissue were taken under ether anesthesia. Nuclear extracts were prepared from liver. Changes in serum AST and ALT activities as well as MDA, TAS, and TOS levels showed that CCl₄ caused lipid peroxidation and liver damage. However, lipid peroxidation and liver damage were reduced in the N-acetyl cysteine group. Increased levels in 8-hydroxy-2-deoxy guanosine and histone acetyltransferase activities, decreased histone deacetylase activities, and DNA breakage detected in nuclear extracts showed that CCl₄ intoxication induces oxidative stress and apoptosis in rat liver. The results of the present study indicate that N-acetyl cysteine has a protective effect on CCl₄-induced DNA damage.     

N-Acetyl Aspartic Acid (NAA) and N-Acetyl Aspartylglutamic Acid (NAAG) in Human Ventricular,Subarachnoid, and Lumbar Cerebrospinal Fluid

N-Acetylaspartic and N-acetylaspartylglutamic acid concentrations in human ventricular, subarachnoid and lumbar cerebrospinal fluid were measured by combined gas chromatography-mass spectrometry using selected ion monitoring with deuterated internal standards. N-Acetylaspartate concentrations were in the range 55, 9, and 1 M, respectively; N-acetylaspartylglutamate concentrations in the same fluids were in the range 8, 3 and 4 M, respectively. There did not appear to be any difference in lumbar fluid concentrations of either compound between control subjects, schizophrenic patients, Alzheimer's disease patients and a pooled group of patients with neurological degeneration. Ventricular concentrations of both compounds were greatly increased in deceased patients suggesting that maintenance of their intracellular concentrations is probably energy dependent. The concentrations of these compounds in lumbar cerebrospinal fluid from living, and ventricular cerebrospinal fluid from deceased subjects were weakly correlated with one another. In lumbar fluid neither compound appeared to be correlated with age. Analysis of serially collected lumbar samples from two subjects showed a weak concentration gradient for both compounds. Neither antipsychotic medication nor the acid transport inhibitor probenecid had any effect on lumbar concentrations of either compound. Attempts to use anion exchange high pressure liquid chromatography with UV detection for measurement of the low concentrations of N-acetylaspartate found in cerebrospinal fluid from living subjects were unsuccessful.     

Surface-Bonded Antimicrobial Activity of an Organosilicon Quaternary Ammonium Chloride

The hydrolysis product of 3-(trimethoxysilyl)-propyldimethyloctadecyl ammonium chloride exhibited antimicrobial activity against a broad range of microorganisms while chemically bonded to a variety of surfaces. The chemical was not removed from surfaces by repeated washing with water, and its antimicrobial activity could not be attributed to a slow release of the chemical, but rather to the surface-bonded chemical.     

Detoxification rates of wild herbivorous woodrats (Neotoma)

The detoxification systems of mammalian herbivores are thought to have evolved in response to the ingestion of plant secondary compounds. Specialist herbivores consume high quantities of secondary compounds and are predicted to have faster rates of Phase 1 detoxification compared to generalist herbivores. We tested this hypothesis by comparing the performances of a specialist (Neotoma fuscipes) and generalist (Neotoma lepida) herbivore using hypnotic state assays. Herbivores foraging in nature were live trapped and injected with hexobarbital (100 mg/kg). We measured the length of time in the hypnotic state as the time in which the animal was unable to right itself twice in 30 s. The specialist metabolized hexobarbital 1.7 times faster than the generalist (F(1, 19) = 9.31, P = 0.007) as revealed by its significantly shorter time spent in the hypnotic state (56+/-9 min vs. 87+/-8 min, respectively). The results are consistent with the hypothesis that specialists have faster rates of Phase 1 detoxification. This is the first evaluation of the detoxification capability of mammalian herbivores foraging under natural conditions. Hypnotic state assays have broad potential applications to the study of vertebrate-plant interactions.     

Effect of Ammonium Chloride and Methionine Sulfoximine on the Acetylene Reduction of Detached Root Nodules of Peas (Pisum sativum)

Acetylene-reducing activity of detached pea nodules was determined by submerging the nodules in buffer solution [tris(hydroxymethyl)aminomethane-hydrochloride, pH 7.4] containing 100 mM sodium succinate and incubating under a gas phase of 90% O₂ and 10% C₂H₂. The nitrogenase activity was 4 to 8 μmol of C₂H₄ formed per g of nodule fresh weight per h and remained constant for at least 4 h. Addition of NH₄Cl to the buffer solution (at a concentration of 10 mM or more) resulted in a significant decrease of nitrogenase activity, which was more pronounced at higher concentrations of ammonium chloride. The inhibition of nitrogenase activity by NH₄Cl was reversible; when the NH₄Cl-containing buffer solution was replaced by buffer without NH₄Cl, the original activity was partly restored. Treatment of the nodules with NH₄Cl had almost no effect on the amount of nitrogenase, as measured by the acetylene-reducing activity of ethyl-enediaminetetraacetate-toluene-treated bacteroid suspensions. The effect of NH₄Cl was largely eliminated by simultaneous addition of 10 mM methionine sulfoximine to the assay solution. This suggests that the assimilation of ammonium ions by glutamine synthetase controls the functioning of nitrogenase activity in the nodules. However, no effect of glutamine, glutamate, or aspartate on the acetylene reduction by detached nodules could be detected.     

Detoxification of lipopolysaccharide (LPS) by egg white lysozyme

Abstract Recent studies carried out by our group suggest that lysozyme binds to bacterial lipopolysaccharide with a high affinity to produce a complex, and inhibits various biological activities of lipopolysaccharide. Although the basic structure of lipopolysaccharide is independent of the species and strains of Gram-negative bacteria, many structural factors such as O-antigenic polysaccharide, lipid A, substituted groups, and associated molecules, affect the biological activities of lipopolysaccharide. In this study, we prepared lysozyme/lipopolysaccharide complexes using various structures of lipopolysaccharide and compared the activity and physiochemical properties. Native and dansylated lysozyme were found to bind to all tested lipopolysaccharides. The mitogenic activity and TNF production by all tested lipopolysaccharides were significantly reduced by complex formation in vitro. Administration of the complex prepared by various lipopolysaccharides produced significantly less quantities of TNF in the septic shock model. These results suggested that binding of lysozyme to lipopolysaccharide is important for the host both in pathophysiological responses to lipopolysaccharides and in the modification of lipopolysaccharide biological activity.     

Detoxification of acidic catalyzed hydrolysate of Kappaphycus alvarezii (cottonii)

Red seaweed, Kappaphycus alvarezii, holds great promise for use in biofuel production due to its high carbohydrate content. In this study, we investigated the effect of fermentation inhibitors to the K. alvarezii hydrolysate on cell growth and ethanol fermentation. In addition, detoxification of fermentation inhibitors was performed to decrease the fermentation inhibitory effect. 5-Hydroxymethylfurfural and levulinic acid, which are liberated from acidic hydrolysis, was also observed in the hydrolysate of K. alvarezii. These compounds inhibited ethanol fermentation. In order to remove these inhibitors, activated charcoal and calcium hydroxide were introduced. The efficiency of activated charcoals was examined and over-liming was used to remove the inhibitors. Activated charcoal was found to be more effective than calcium hydroxide to remove the inhibitors. Detoxification by activated charcoal strongly improved the fermentability of dilute acid hydrolysate in the production of bioethanol from K. alvarezii with Saccharomyces cerevisiae. The optimal detoxifying conditions were found to be below an activated charcoal concentration of 5%.     

Detoxification of dissolved SO2 (bisulfite) by terricolous mosses

BACKGROUND AND AIMS: The widespread calcifuge moss Pleurozium schreberi is moderately tolerant of SO2, whereas Rhytidiadelphus triquetrus is limited to calcareous soils in regions of the UK that were strongly affected by SO2 pollution in the 20th century. The proposition that tolerance of SO2 by these terricolous mosses depends on metabolic detoxification of dissolved bisulfite was investigated. METHODS: The capacities of the two mosses to accelerate loss of bisulfite from aqueous solutions of NaHSO3 were studied using DTNB [5, 5-dithio-(2-nitrobenzoic acid)] to assay bisulfite, and HPLC to assay sulfate in the incubation solutions. Incubations were performed for different durations, in the presence and absence of light, at a range of solution pH values, in the presence of metabolic inhibitors and with altered moss apoplastic Ca2+ and Fe3+ levels. KEY RESULTS: Bisulfite disappearance was markedly stimulated in the light and twice as great for R. triquetrus as for P. schreberi. DCMU, an inhibitor of photosynthetic electron chain transport, significantly reduced bisulfite loss. CONCLUSIONS: Bisulfite (SO2) tolerance in these terricolous mosses involves extracellular oxidation using metabolic (photo-oxidative) energy, passive oxidation by adsorbed Fe3+ (only available to the calcifuge) and probably also internal metabolic detoxification.     

Apoptosis: a process with a (beta)NAC for complexity

Most programmed cell deaths in the nematode C. elegans require ced-3 caspase activity. In a recent paper, reveal a new C. elegans death inhibitor, icd-1, whose loss can promote apoptosis independently of ced-3.     

Activity of Triclopyr Herbicide Enhanced by Combination with Cobalt Chloride or Ammonium Nitrate

Cobalt chloride at a rate of 16 oz/acre when combined with the auxin herbicide triclopyr at 1 oz/acre synergistically enhanced the activity of the herbicide. A similar response was elicited by the addition of ammonium nitrate. Mg²⁺ or Mn²⁺ was antagonistic. The enhancement of activity by Co²⁺ or NH⁺ ₄ appeared not to be the result of a simple enhancement of triclopyr uptake by the cations. Triclopyr uptake, as determined by gas chromatography/microwave-induced plasma/chemical reaction interface mass spectroscopy, was unaffected by the cation additions. Received March 31, 1997; accepted March 24, 1998     

Cysteine pK(a) values for the bacterial peroxiredoxin AhpC

Salmonella typhimurium AhpC is a founding member of the peroxiredoxin family, a ubiquitous group of cysteine-based peroxidases with high reactivity toward hydrogen peroxide, organic hydroperoxides, and peroxynitrite. For all of the peroxiredoxins, the catalytic cysteine, referred to as the peroxidatic cysteine (C(P)), acts as a nucleophile in attacking the peroxide substrate, forming a cysteine sulfenic acid at the active site. Because thiolates are far stronger nucleophiles than thiol groups, it is generally accepted that cysteine-based peroxidases should exhibit pK(a) values lower than an unperturbed value of 8.3-8.5. In this investigation, several independent approaches were used to assess the pK(a) of the two cysteinyl residues of AhpC. Methods using two different iodoacetamide derivatives yielded unperturbed pK(a) values (7.9-8.7) for both cysteines, apparently due to reactivity with the wrong conformation of C(P) (i.e., locally unfolded and flipped out of the active site), as supported by X-ray crystallographic analyses. A functional pK(a) of 5.94 +/- 0.10 presumably reflecting the titration of C(P) within the fully folded active site was obtained by measuring AhpC competition with horseradish peroxidase for hydrogen peroxide; this value is quite similar to that obtained by analyzing the pH dependence of the epsilon(240) of wild-type AhpC (5.84 +/- 0.02) and similar to those obtained for two typical 2-cysteine peroxiredoxins from Saccharomyces cerevisiae (5.4 and 6.0). Thus, the pK(a) value of AhpC balances the need for a deprotonated thiol (at pH 7, approximately 90% of the C(P) would be deprotonated) with the fact that thiolates with higher pK(a) values are stronger nucleophiles.