NMR studies on energy metabolism of immobilized primary neurons and astrocytes during hypoxia, ischemia and hypoglycemia

Changes in high-energy phosphate metabolites (ATP and phosphocreatine) were monitored, in real time, by 31P-nuclear magnetic resonance in primary cell cultures of neurons and astrocytes during periods of hypoxia, ischemia and hypoglycemia, and also during the recovery periods following the re-establishment of standard conditions. Cells were immobilized in basement membrane gel threads and perfused with oxygen-depleted medium (oxygen concentration below 30 microM), to create hypoxic conditions, or with aerobic medium (oxygen concentration approximately 460 microM) containing different concentrations of glucose (hypoglycemia). Ischemic conditions were imposed by stopping perfusion for different periods of time (15 min to 2 h). The experimental set-up enabled the acquisition of 31P-spectra with high signal-to-noise ratio within 10-20 min for both cell types. The effect of hypoxia on glucose metabolism was assessed by 13C-NMR using [1-13C]glucose as substrate. The levels of ATP and PCr in astrocytes were unaffected during hypoxia (up to 2 h), but decreased notably under ischemia. In neurons, hypoxic periods caused a sharp drop of the ATP and PCr levels, and considerable damage to the capacity of neurons to replenish the ATP and PCr pools upon returning to normoxic conditions. However, neurons were remarkably less sensitive to ischemic conditions, the ATP and PCr pools being restored quickly, even after 2 h under challenging conditions. The data show that neurons were more resistant to ischemia than astrocytes, and suggest that the capacity to sustain the pools of ATP and PCr was part of the neuronal protective strategy.     

Sequential synthesis and secretion of pectinases by Penicillium frequentans

Penicillium frequentans synthesized eleven polygalacturonases and three pectinesterases when grown in the presence of pectin, sodium polypectate or monogalacturonic acid. When glucose was the sole carbohydrate source in the medium two of these polygalacturonases and one pectinesterase were produced. The enzymes produced under any of these conditions degraded pectic substrates to monogalacturonic acid, suggesting that this monosaccharide or its metabolites should induce the pectinolytic complex. All pectinesterases and most of the extracellular polygalacturonases were synthesized after the 2nd hour of incubation. The pectinases produced by Penicillium frequentans were not secreted at the same time but after 5 hours of incubation all of them could be detected outside the cell those detected only inside the cell were probably membrane-associated or unglycosylated forms of the extracellular pectinases.     

Albumin adsorption on alkanethiols self-assembled monolayers on gold electrodes studied by chronopotentiometry

Chronopotentiometry was used to study the adsorption of human serum albumin (HSA) to self-assembled monolayers with the following terminal functional groups: CH(3), COOH and OH. Surfaces were characterized by X-ray photoelectron spectroscopy, water contact angle measurements and cyclic voltammetry. HSA coverage of the different SAMs was investigated by chronopotentiometry and the total amount of adsorbed protein was determined using radiolabelled albumin. Both techniques have demonstrated that HSA adsorption to the different SAM-modified electrodes increases in the following order: OH相似文献   

Changed accumbal responsiveness to alcohol in rats pre-treated with nicotine or the cannabinoid receptor agonist WIN 55,212-2

Alcohol, nicotine, and cannabinoid acutely increase the activity of the mesolimbic dopamine (DA) pathway. Although polysubstance consumption is a common pattern of abuse in humans, little is known about dopamine release following pre-exposure to these drugs. The purpose of this study was to test whether alcohol-induced dopamine release into the nucleus accumbens (NAc) shell is modified by different pre-treatments: water (i.g.), alcohol (1 g/kg, i.g.), nicotine (0.4 mg/kg, s.c.), and WIN 55,212-2 (1 mg/kg, s.c.). Male Wistar rats were treated (i.g.) for 14 days with either water or alcohol. In the following 5 days rats were injected (s.c.) with vehicle, nicotine, or WIN 55,212-2. Finally, a cannula was surgically implanted into the NAc shell and alcohol-induced extracellular dopamine release was monitored in freely moving rats. Alcohol (1 g/kg; i.g.) only increased the release of dopamine when animals were previously treated with water. This DA increase was markedly inhibited by (subchronic) treatment (5 days) with nicotine or WIN 55-212-2 as well as by previous (chronic) exposure to alcohol (14 days). These data demonstrate that pre-treatment with nicotine and the cannabinoid agonist WIN 55,212-2 is able to change the sensitivity of the NAc shell in response to a moderate dose of alcohol. Therefore, cannabinoid and nicotine exposure may have important implications on the rewarding effects of alcohol, because these drugs lead to long-lasting changes in accumbal dopamine transmission.     

Mutagenicity,antioxidant potential,and antimutagenic activity against hydrogen peroxide of cashew (Anacardium occidentale) apple juice and cajuina

Fresh and processed cashew (Anacardium occidentale) apple juice (CAJ) are among the most popular drinks in Brazil. Besides their nutritional benefits, these juices have antibacterial and antitumor potential. The chemical constituents of both the fresh juice and the processed juice (cajuina) were analyzed and characterized as complex mixtures containing high concentrations of vitamin C, various carotenoids, phenolic compounds, and metals. In the present study, these beverages exhibited direct and rat liver S9-mediated mutagenicity in the Salmonella/microsome assay with strains TA97a, TA98, and TA100, which detect frameshifts and base pair substitution. No mutagenicity was observed with strain TA102, which detects oxidative and alkylating mutagens and active forms of oxygen. Both CAJ and cajuina showed antioxidant activity as determined by a total radical-trapping potential assay. To test whether this antioxidant potential might result in antimutagenesis, we used a variation of the Salmonella/microsome assay that included pre-, co-, and posttreatment of hydrogen peroxide-exposed Salmonella typhimurium strain TA102 with the juices. CAJ and cajuina protected strain TA102 against mutation by oxidative damage in co- and posttreatments. The antimutagenic effects during cotreatment with hydrogen peroxide may be due to scavenging free radicals and complexing extracellular mutagenic compounds. The protective effects in posttreatment may be due to stimulation of repair and/or reversion of DNA damage. The results indicate that CAJ and cajuina have mutagenic, radical-trapping, antimutagenic, and comutagenic activity and that these properties can be related to the chemical constituents of the juices.     

Evaluation of Etest strips for rapid susceptibility testing of Mycobacterium tuberculosis

In this study, our objective was to evaluate Etest strips containing exponential gradients of isoniazid (INH), rifampin (RIF), and streptomycin (STR) for susceptibility testing of Mycobacterium tuberculosis. M. tuberculosis isolates were tested for antimicrobial susceptibilities by the standard proportion method using L?wenstein-Jensen (LJ) medium and by the Etest. The MICs determined by the Etest were obtained at 5, 7, or 10 days. In some strains with Etest-discrepant results, radiometric susceptibility testing (BACTEC) was performed to determine a consensus result. M. tuberculosis concordance between the two methods was 97% (86 of 89 isolates) for RIF, 96% for INH (84 of 87 isolates), and 80% (61 of 76 isolates) for STR. Most of the MICs determined by the Etest were easy to interpret and readable within 5 days. Results correlated well with those obtained by the LJ proportion and BACTEC methods for INH and RIF. However, a high proportion of false-sensitive and false-resistant results were observed, most often for STR. We also observed that variations in the inoculum size of M. tuberculosis isolates affected the MICs to a substantial degree. These discrepancies, along with the expense of the media, the Etest strips, and the specialized equipment required (CO2 incubator), make this method less useful in developing countries.     

Adenocarcinomas of the nasal cavity and paranasal sinuses: a clinicopathological and immunohistochemical study of 14 cases

Aims : To evaluate the clinicopathological profile of 14 cases of nasal and paranasal sinusal adenocarcinoma, and to assess the usefulness of immunohistochemistry in the differential diagnosis of primary and metastatic intestinal-type adenocarcinoma.
Methods and results : Fourteen cases of nasal and paranasal adenocarcinoma, treated at IPOFG, Lisbon, between 1976 and 2002, were studied. Clinical records were reviewed and expression of cytokeratin (CK)7 and CK20 and of neuroendocrine markers was evaluated. The male : female ratio was 3 : 1, and the mean age of the patients was 65.3 years. Ten cases occurred in the paranasal sinuses. There was a history of professional exposure to dust in three patients. Twelve cases were high-grade intestinal type adenocarcinomas (ITAC) and two were low-grade. CK7 was present in 2/9 ITAC cases and CK20 in 8/9 ITAC and in cases of mixed and mucinous histology. All high-grade cases showed neuroendocrine differentiation. Seven of the 12 patients with high-grade adenocarcinoma died of the disease, with a mean follow-up of 47.4 months.
Conclusions : Nasal and paranasal adenocarcinoma mostly occurs in men in the 7th decade. ITAC is the most frequent histological type. The pattern of CK7/CK20 was not useful in the distinction between primary and metastatic intestinal adenocarcinoma. However, in the former, neuroendocrine differentiation proved to be a valuable tool in that distinction.     

Comparison of protective immunity elicited by recombinant vaccinia viruses that synthesize E or NS1 of Japanese encephalitis virus.

Immunization with recombinant vaccinia viruses that specified the synthesis of Japanese encephalitis virus (JEV) glycoproteins protected mice from a lethal intraperitoneal challenge with JEV. Recombinants which coexpressed the genes for the structural glycoproteins, prM and E, elicited high levels of neutralizing (NEUT) and hemagglutination inhibiting (HAI) antibodies in mice and protected mice from a lethal challenge by JEV. Recombinants expressing only the gene for the nonstructural glycoprotein, NS1, induced antibodies to NS1 but provided low levels of protection from a similar challenge dose of JEV. Antibodies to the NS3 protein in postchallenge sera, representing the degree of infection with challenge virus, were inversely correlated to NEUT and HAI titers and levels of protection. These results indicate that although vaccinia recombinants expressing NS1 can provide some protection from lethal JEV infection, recombinants expressing prM and E elicited higher levels of protective immunity.     

Increased serum levels of growth hormone and insulin-like growth factor-I associated with simultaneous decrease of circulating insulin in postmenopausal women receiving hormone replacement therapy

OBJECTIVE: Decreases in circulating growth hormone (GH) and its main biological messenger insulin-like growth factor-I (IGF-I) have been interpreted as part of the aging process. Because estrogens participate in modulating GH synthesis and secretion, hypoestrogenism in menopausal women may lead to GH deficiency. The aim of the present study was to determine the effect of hormone replacement therapy (HRT) on both GH and IGF-I levels as well as insulin concentrations in 50 menopausal women. DESIGN: Patients were assigned randomly into two treatment groups of 25 each; one group received three cycles of conjugated equine estrogen (CEE) 0.625 mg/day for 21 days, and the other, 1.25 mg/day during 21 days. Each also received chlormadinone acetate for 5 days. There was a control group consisting of regularly menstruating women. RESULTS: In the menopausal women, HRT increased significantly (p < 0.001) the low levels of GH and IGF-I; on the contrary the baseline insulin levels declined (p < 0.001) with HRT. A significant linear correlation (r = 0.90) was found between GH and IGF-I as well as with estradiol levels (r = 0.74) in the group of menopausal women receiving CEE 0.625 mg/day. This group of patients had a significant correlation (r = -0.63) between insulin and estradiol levels. No correlation was observed in the group receiving CEE 1.25 mg/day. CONCLUSIONS: HRT restored GH, IGF-I, and insulin levels to normal values in all women. Further research needs to be done to establish the beneficial effect of HRT regarding the prevention of the metabolic effects presumably caused by derangement in the somatotropic axis associated with aging.