Glutathione peroxidase overexpression inhibits cytochrome C release and proapoptotic mediators to protect neurons from experimental stroke.

BACKGROUND AND PURPOSE: Ischemic injury and reperfusion increases superoxide (O2-) production and reduces the ability of neurons to scavenge free radicals, leading to the release of cytochrome c and apoptosis. Here we test whether overexpression with the use of gene therapy of the antioxidant glutathione peroxidase (Gpx), delivered before or after experimental stroke, is protective against ischemic injury. METHODS: Sixty-two rats underwent middle cerebral artery occlusion for 1 hour. Defective herpes simplex viral vectors expressing Gpx/lacZ or lacZ alone (control) were delivered into each striatum 12 hours before or 2 or 5 hours after ischemia onset. RESULTS: Striatal neuron survival at 2 days was improved by 36% when Gpx was delivered 12 hours before ischemia onset, 26% with a 2-hour delay, and 25% when delayed 5 hours. After ischemia, Gpx overexpression significantly reduced cytosolic translocation of cytochrome c and increased the proportion of Bcl-2-positive cells compared with cells transfected with control vector. Bax and activated caspase-3, while present in control-transfected neurons after ischemia, were rarely noted in Gpx-transfected cells. CONCLUSIONS: Expression from these herpes simplex viral vectors begins 4 to 6 hours after injection, which suggests a 9- to 11-hour temporal therapeutic window for Gpx. This is the first study to show that overexpression of Gpx with the use of gene therapy protects against experimental stroke, even with postischemic transfection, and the neuroprotective mechanism involves attenuation of apoptosis-related events.

Review article: Nutritional therapy in alcoholic liver disease.

Chronic alcohol consumption may lead to primary and secondary malnutrition. In particular, protein energy malnutrition not only aggravates alcoholic liver disease but also correlates with impaired liver function and increased mortality. Therefore, in these patients, adequate nutritional support should be implemented in order to improve their prognosis. Clinical trials addressing this issue have shown that nutritional therapy either enterally or parenterally improves various aspects of malnutrition, and there is increasing evidence that it may also improve survival. Therefore, malnourished alcoholics should be administered a diet rich in carbohydrate- and protein-derived calories preferentially via the oral or enteral route. Micronutrient deficiencies typically encountered in alcoholics, such as for thiamine and folate, require specific supplementation. Patients with hepatic encephalopathy may be treated with branched-chain amino acids in order to achieve a positive nitrogen balance. Fatty liver represents the early stage of alcoholic liver disease, which is usually reversible with abstinence. Metadoxine appears to improve fatty liver but confirmatory studies are necessary. S-adenosyl-L-methionine may be helpful for patients with severe alcoholic liver damage, since various mechanisms of alcohol-related hepatotoxicity are counteracted with this essential methyl group donor, while a recent large trial showed that the use of polyenylphosphatidylcholine is of limited efficacy.

VEGF mRNA expressed in microvessels of neonatal and adult rat cerebral cortex.

We measured mRNA levels of vascular endothelial growth factor (VEGF) and its Flk-1/KDR receptor in isolated cerebral cortical microvessels and in the cerebral cortex of neonatal (1 week) and adult (11 week) rats using reverse transcription-polymerase chain reaction (RT-PCR). Cerebral microvessels were isolated by density centrifugation, mesh filtration and passage through glass bead columns. The dominant cell types in this preparation are endothelial cells and pericytes. Among the four isoforms of VEGF mRNA expressed in these tissues, VEGF(165) was dominant (67% higher than VEGF(189) or VEGF(206)). All isoforms of VEGF were higher in adult cortical microvessels than in cortical homogenates. In isolated microvessels, VEGF mRNA for all isoforms combined was 70% higher in the neonate than in the adult. VEGF receptor Flk-1/KDR mRNA was also present in cortical microvessels and was higher in neonatal than in adult microvessels. The results suggest that VEGF is normally expressed in cerebral microvessels of both neonates and adults. Whether the source of VEGF is the endothelial cell or pericyte, will determine if VEGF has autocrine or paracrine actions. The results also support the hypothesis that microvascular cell turnover continues in the adult brain.

Overexpression of HSP72 after induction of experimental stroke protects neurons from ischemic damage.

The 72-kD inducible heat shock protein (HSP72) can attenuate cerebral ischemic injury when overexpressed before ischemia onset. Whether HSP72 overexpression is protective when applied after ischemia onset is not known, but would have important clinical implications. Fifty-seven rats underwent middle cerebral artery occlusion for 1 hour. Defective herpes simplex viral (HSV) vectors expressing hsp72 with lacZ as a reporter were delivered 0.5, 2, and 5 hours after ischemia onset into each striatum. Control animals received an identical vector containing only lacZ. Striatal neuron survival at 2 days was improved by 23% and 15% when HSP72 vectors were delayed 0.5 and 2 hours after ischemic onset, respectively ( P < 0.05). However, when delayed by 5 hours, HSP72 overexpression was no longer protective. This is the first demonstration that HSP72 gene transfer even after ischemia onset is neuroprotective. Because expression from these HSV vectors begins 4 to 6 hours after injection, this suggests that the temporal therapeutic window for HSP72 is at least 6 hours after ischemia onset. Future strategies aimed at enhancing HSP72 expression after clinical stroke may be worth pursuing. The authors suggest that in the future HSP72 may be an effective treatment for stroke.

New challenges for nursing informatics.

Long a leader in health informatics, nursing faces new challenges. The full and effective use of technology requires an understanding of cognitive processes and organizational behavior. Nursing can play a key role in addressing aims supportive of a new vision of health informatics. The evolving paradigm for knowledge transfer will give rise to new educational models and new institutional entities which will nurture learning and relearning.

Supporting collaboration through a nursing informatics curriculum stage II.

Collaboration is at the center of the process used to design, implement and evaluate an integrated informatics curriculum in a baccalaureate nursing program. This paper describes the second stage of a process to design the informatics nursing courses. The challenges to foster faculty collaborative relationships as well as to enhance the course content of all nursing informatics curriculum. A number of strategies were used to develop the collaborative efforts between the faculty and nursing staff in the clinical agencies. Information technology was incorporated into the didactic and clinical portions of courses through the use of creative teaching strategies. Therefore, the faculty have ensured a blend of information, technology, and the clinical care process throughout the curriculum.

An integrated informatics curriculum in a baccalaureate nursing program.

As health care requirements change, nurses will not only have to process and communicate more information, but the nature and types of this information will dramatically change as well. It is imperative that nurses understand the potential information technologies offer to assist the nurse in this expanded role. This paper describes an innovative endeavor to incorporate information technology with its undergraduate nursing program. The challenge was to design a program that would help develop the students' skills to critically appraise their information needs and conceptually evaluate the utility of gathering information in providing patient care. After completing the first nursing informatics course, there was an increase in the students' perception and understanding of the uses of information technology to support the nursing process in providing patient care.

Apparent high degree of asymmetry of protein arrangement in the Escherichia coli outer cell envelope membrane.

Ghosts from Escherichia coli have been oxidized with CuSO4-o-phenanthroline or ferricyanide-ferrocene. Upon oxidation they became resistant to boiling dodecyl sulfate. The resulting rod-shaped "oxidation containers" apparently held together by disulfide bridges, are practically pure protein. They are soluble in dodecyl sulfate when reduced and they contain a set of about 30 different polypeptide chains. The four major ghost membrane proteins are not represented among the "oxidation proteins." Comparison of data obtained from digestion of ghosts with trypsin or particle-bound trypsin showed that most of the "oxidation proteins" appear to be located at the outer surface of the ghost membrane which is derived from the outer cell envelope membrane. One of the major ghost membrane proteins, II, is partially digested by trypsin, and it is shown that its trypsin sensitive part is also exposed only at the outer surface of the ghost membrane. Native cells could be oxidized only with low yields of "oxidation containers." However, cell envelopes prepared without detergents or chelating agents, as well as cells depleted of phospholipid or treated with sucrose-Triton X-100, are completely accessible to oxidation. In each case, the same set of proteins as that present in "oxidation containers" from ghosts was found to be covalently linked. Treatment of cells with trypsin caused the loss of about five "oxidation proteins" and a complete loss of oxidizability of the ghosts derived from these cells. It therefore appears that arrangement and localization of the "oxidation proteins" are not greatly different in cells and in ghosts, i.e., that these proteins are also situated asymmetrically at the outer cell envelope membrane.

Cell envelope and shape of Escherichia coli K12.

Rod-shaped "ghosts" that are free of murein have been isolated from E. coli. The shape of these "ghosts" is maintained by a unit membrane soluble in sodium dodecyl sulfate. Ghosts consist of about 20-30% phospholipid (almost exclusively phosphatidylethanolamine) and 50-60% protein; a large fraction of the remaining material is lipopolysaccharide. Sodium dodecyl sulfate-gel electrophoresis reveals 4-5 different bands corresponding to molecular weights between 10,000 and 40,000. Treatment of ghosts with Pronase reduces this number to 3, and the rod shape still is not lost. Results of treatment of ghosts with a crude extract from Dictyostelium discoideum have supplied tentative evidence that at least one of these proteins is involved in the maintenance of rod shape. It does not appear too unlikely that these polypeptide chains are the final products of the genetic information specifying cellular shape.

Molecular structure of the pyruvate dehydrogenase complex from Escherichia coli K-12.

The pyruvate dehydrogenase core complex from E. coli K-12, defined as the multienzyme complex that can be obtained with a unique polypeptide chain composition, has a molecular weight of 3.75 x 10(6). All results obtained agree with the following numerology. The core complex consists of 48 polypeptide chains. There are 16 chains (molecular weight = 100,000) of the pyruvate dehydrogenase component, 16 chains (molecular weight = 80,000) of the dihydrolipoamide dehydrogenase component, and 16 chains (molecular weight = 56,000) of the dihydrolipoamide dehydrogenase component. Usually, but not always, pyruvate dehydrogenase complex is produced in vivo containing at least 2-3 mol more of dimers of the pyruvate dehydrogenase component than the stoichiometric ratio with respect to the core complex. This "excess" component is bound differently than are the eight dimers in the core complex.