NAP prevents hippocampal oxidative damage in neonatal rats subjected to hypoxia-induced seizures

Neonatal seizures in which hypoxic-ischemic encephalopathy is the main triggering etiology have a challenging diagnosis and limited efficacy of treatment. NAP (NAPVSIPQ) has shown extensive neuroprotective and antioxidant capacity in vitro and in vivo. To evaluate its neuroprotective role in the context of seizures associated with perinatal hypoxia, we assessed the integrity of DNA and lipid membranes as well as the redox status in the hippocampus of 10-day-old rats exposed to hypoxia-induced seizures (HS) with and without NAP treatment. Rats were exposed to transient global hypoxia (12 min exposure to 5–7% O₂ was able to induce electrographic seizures) or room air with subsequent intraperitoneal NAP (0.03, 0.3 or 3 μg/g) or vehicle administration. Results showed elevated DNA damage immediately after the insult until 72 h post-HS, while oxidized bases were only detected 3, 6 and 24 h later. In addition, thiobarbituric acid reactive species peaked at 6 h in parallel with decreased levels of reduced glutathione between 3 and 72 h post-HS insult. Our findings expand on the knowledge about the time course of HS-induced oxidative damage and demonstrate for the first time that a single NAP injection dose-dependently prevents HS-induced oxidative damage to DNA and lipid membranes, in correlation with modulation of the glutathione system. Hence, NAP may represent a promising therapeutic strategy for avoiding HS-induced oxidative damage.     

Localization of phospholipase C beta 1 on the detergent-resistant membrane microdomain prepared from the synaptic plasma membrane fraction of rat brain

The membrane microdomain (MD), such as detergent-resistant low-density membrane microdomain fraction (DRM), has been paid much attention because many signal-transducing molecules are recovered in this fraction, although precise localization and interactions of these molecules are largely unclear. To identify neuronal MD-localized proteins, monoclonal antibodies (mAbs) against the DRM-components of synaptic plasma membrane fraction (SPM) were produced and the antigens were characterized. One of the antigens reacted with two closely positioned bands of about 140 kDa in SDS-PAGE and the antigen showed age-dependent localization on DRM. The antigen was immunoprecipitated with the mAb after partial solubilization with 0.6 M NaCl from SPM-derived DRM and identified as phospholipase C beta 1 through mass analysis. The identity was further confirmed with Western blotting using a specific polyclonal antibody. The enzyme purified from the DRM was activated by the alpha subunit of trimeric G protein, Gq, expressed in HEK293 cells. The lipid composition of the liposomes affected the enzymatic activity and the addition of NAP-22, a neuronal DRM-localized protein, inhibited the activity. These results suggest that there exists a signal-transducing MD that performs important roles in neuronal functions through PIP(2) signaling and Ca(2+) mobilization.     

Novel therapeutics based on tau/microtubule dynamics: WO2008084483

Background: The present patent deals with the generation of peptides derived from the activity-dependent peptide and tau mimetic to study its effect on microtubule stability, its ability to bind to tubulin and MAPs, as well as promoting cell survival. Objective: To analyze these peptides and their effects as potential therapeutic elements for neurodegenerative diseases. Methods: We review the action of the peptides described by Gozes and collaborators and compare the effectiveness with those already reported in the literature for Alzheimer's disease. Conclusion: The research of Dr. Gozes and collaborators has shown that the addition of picomolar concentration of the peptides promotes cell survival, by interacting with tubulin and stabilizing the microtubules. Based on the results, these peptides seem to be very attractive candidates for therapeutical intervention in neurodegenerative diseases.     

Characterization of 6α- and 6β-N-heterocyclic substituted naltrexamine derivatives as novel leads to development of mu opioid receptor selective antagonists

As important pharmacological probes, highly selective opioid receptor antagonists are essential in opioid receptor structural characterization and opioid agonist functional studies. At present, a nonpeptidyl, highly selective, and reversible mu opioid receptor antagonist is still not available. Among a series of novel naltrexamine derivatives that have been designed and synthesized following molecular modeling studies, two compounds, NAP and NAQ, were identified as leads based on the results of in vitro and in vivo pharmacological assays. Both of them displayed high binding affinity and selectivity to the mu opioid receptor. Further pharmacokinetic and functional characterization revealed that NAP seems to be a peripheral nervous system agent while NAQ seems to be a central one. Such characteristics provide two distinguished potential application routes for these two agents and their derivatives. These results also supported our hypothesis that they may serve as leads to develop more potent and selective antagonists for the mu opioid receptor.     

CRP、NAP、WBC、DC的联合检测在中晚期肿瘤患者细菌感染中的诊断价值

目的比较和评价C反应蛋白(CRP)、中性粒细胞碱性磷酸酶染色(NAP)、白细胞计数(WBC)及分类计数(DC)四项指标的联合检测在中晚期肿瘤患者细菌感染中的临床诊断价值。方法对104例中晚期肿瘤患者其中明确感染者40例,无感染者64例,行CRP、NAP、WBC、DC四项指标的联合检测。结果CRP在两组肿瘤患者中均明显升高,感染组的四项指标均高于非感染组,均有显著的统计学差异。结论CRP、NAP、WBC、DC四项指标的联合检测有利于中晚期肿瘤患者细菌感染的诊断,有利于及早控制中晚期肿瘤患者的细菌感染和避免不必要的抗生素应用。     

血涂片与骨髓涂片碱性磷酸酶染色积分比较

目的中性粒细胞碱性磷酸酶（NAP）染色常规使用血片或骨髓片。比较血片与骨髓片NAP积分差异，以指导实践。方法42例患者。取其血片及骨髓片各1张，同时进行NAP染色。结果血片NAP积分均数59.38稍高于骨髓片NAP积分均数49.19；两者阳性率呈正相关（rs=0.8091，P〈0.01）；两者积分呈正相关（rs=0.7977，P〈0.01）。结论血片及骨髓片可相互代替或同时进行NAP染色。     

Nitric oxide inhibits insulin-degrading enzyme activity and function through S-nitrosylation

Insulin-degrading enzyme (IDE) is responsible for the degradation of a number of hormones and peptides, including insulin and amyloid β (Aβ). Genetic studies have linked IDE to both type 2 diabetes and Alzheimer's disease. Despite its potential importance in these diseases, relatively little is known about the factors that regulate the activity and function of IDE. Protein S-nitrosylation is now recognized as a redox-dependent, cGMP-independent signaling component that mediates a variety of actions of nitric oxide (NO). Here we describe a mechanism of inactivation of IDE by NO. NO donors decreased both insulin and Aβ degrading activities of IDE. Insulin-degrading activity appeared more sensitive to NO inhibition than Aβ degrading activity. IDE-mediated regulation of proteasome activity was affected similarly to insulin-degrading activity. We found IDE to be nitrosylated in the presence of NO donors compared to that of untreated enzyme and the control compound. S-nitrosylation of IDE enzyme did not affect the insulin degradation products produced by the enzyme, nor did NO affect insulin binding to IDE as determined by cross-linking studies. Kinetic analysis of NO inhibition of IDE confirmed that the inhibition was noncompetitive. These data suggest a possible reversible mechanism by which inhibition of IDE under conditions of nitrosative stress could contribute to pathological disease conditions such as Alzheimer's disease and type 2 diabetes.     

A single administration of the peptide NAP induces long-term protective changes against the consequences of head injury

The femtomolar-acting eight-amino-acid peptide (NAP), derived from activity-dependent neuroprotective protein (ADNP), provides long-term protection against the deleterious effects of closed head injury (CHI) in mice. Fifteen minutes after injury, mice were divided into two groups, control and NAP-treated and a single subcutaneous injection of NAP or vehicle was administered. A third group served as sham-treated (not subjected to head trauma). Each mouse was assessed for its clinical function, using neurological severity score, at various time intervals following CHI, up to 30–45 d. Total cerebral cortex RNA was prepared from the site of injury of CHI mice, and from parallel regions in peptide-treated and sham brains. RNA was then reversed transcribed to yield radioactive cDNA preparations that were hybridized to Atlas array membranes containing 1200 cDNAs spots. Comparison of sham-treated individual mice showed differential expression levels of at least 15 mRNA species. Furthermore, results indicated that one of the genes that did not change among individuals but specifically increased after CHI and decreased after NAP treatment was the cell surface glycoprotein Mac-1 (CD11B antigen). Thus, Mac-1 is suggested as a marker for the long-term outcome of head injury and as a potential target for NAP protective actions.     

Prolonged post-tetanic depolarization of frog dorsal root fibers

In the isolated frog spinal cord, tetanic stimulation of a dorsal root resulted in a sustained negative potential recorded from that root. This negative potential was followed by a large (5–12 mV) and slow (5–60 s) negative after-potential which persisted after the train of impulses and which was succeeded by a long-lasting positive potential. The negative after-potential was postulated to be largely a consequence of excess extracellular K⁺ accumulated during the tetanus, because the negative after-potential and the extracellular [K⁺] measured with K⁺-sensitive microelectrodes were a function of the stimulating current, the stimulation frequency and the duration of the tetanus. Both measures were augmented by procedures which interfered with the clearance of K⁺ from the extracellular space (addition of dinitrophenol or ouabain, partial substitution of Li⁺ for Na⁺, elimination of K⁺from the superfusate and cooling). The negative after-potential and the liberation of much of the excess extracellular K⁺ were reduced by increasing the superfusing divalent ion concentration with 4mM Ca²⁺or 50–100 μM Mn²⁺. Both measures were augmented in low Ca²⁺-containing Ringer's solution. The results with low concentrations of Ca²⁺, Na⁺ and K⁺ and with high concentrations of Ca²⁺ indicate that the negative after-potential was not caused by a conductance increase for these cations. Reduction of extracellular [Cl^?] did not change the negative after-potential making it unlikely that this anion is a factor in its production. The negative after-potential was not mediated by γ-aminobutyrate because its amplitude and duration were increased by the γ-aminobutyrate antagonist picrotoxin, and were reduced by pentobarbital which enhances the actions of γ-aminobutyrate.Application of the excitatory amino acid antagonistsdl-α-aminoadipic acid and glutamic acid diethylester eliminated the negative after-potential and markedly reduced the amount of K⁺ liberated by a tetanus. These results suggest that the K⁺ release that produces the negative after-potential may result from the action of an excitatory amino acid such asl-glutamate orl-aspartate either synaptically released or released from nonsynaptic regions of afferent fibers.