Aromatase expression and cell proliferation following injury of the adult zebra finch hippocampus

Estrogens can be neuroprotective following traumatic brain injury. Immediately after trauma to the zebra finch hippocampus, the estrogen-synthetic enzyme aromatase is rapidly upregulated in astrocytes and radial glia around the lesion site. Brain injury also induces high levels of cell proliferation. Estrogens promote neuronal differentiation, migration, and survival naturally in the avian brain. We suspect that glia are a source of estrogens promoting cell proliferation after neural injury. To explore this hypothesis, we examined the spatial and temporal relationship between glial aromatase expression and cell proliferation after neural injury in adult female zebra finches. Birds were ovariectomized and given a blank implant or one filled with estradiol; some birds were also administered an aromatase inhibitor or vehicle. All birds received penetrating injuries to the right hippocampus. Twenty-four hours after lesioning, birds were injected once with BrdU to label mitotically active cells and euthanized 2 h, 24 h, or 7 days later. The brains were processed for double-label BrdU and aromatase immunocytochemistry. Injury-induced glial aromatase expression was unaffected by survival time and aromatase inhibition. BrdU labeling was significantly reduced at 24 h by ovariectomy and by aromatase inhibition; effects were partially reversed by E2 replacement. Irrespective of ovariectomy, the densities of aromatase immunoreactive astrocytes and BrdU-labeled cells at known distances from the lesion site were highly correlated. These data suggest that injury-induced glial aromatization may influence the reorganization of injured tissue by providing a rich estrogenic environment available to influence cellular incorporation.     

Adhesive thermosensitive gels for local delivery of viral vectors

Local delivery of viral vectors can enhance the efficacy of therapies by selectively affecting necessary tissues and reducing the required vector dose. Pluronic F127 is a thermosensitive polymer that undergoes a solution–gelation (sol–gel) transition as temperature increases and can deliver vectors without damaging them. While pluronics can be spread over large areas, such as the surface of an organ, before gelation, they lack sufficient adhesivity to remain attached to some tissues, such as the surface of the heart or mucosal surfaces. Here, we utilized blends of pluronic F127 and polycarbophil (PCB), a mucoadhesive agent, to provide the necessary adhesivity for local delivery of viral vectors to the cardiac muscle. The effects of PCB concentration on adhesive properties, sol–gel temperature transition and cytocompatibility were evaluated. Rheological studies showed that PCB decreased the sol–gel transition temperature at concentrations >1% and increased the adhesive properties of the gel. Furthermore, these gels were able to deliver viral vectors and transduce cells in vitro and in vivo in a neonatal mouse apical resection model. These gels could be a useful platform for delivering viral vectors over the surface of organs where increased adhesivity is required.     

Sonochemically fabricated microelectrode arrays for biosensors offering widespread applicability: Part I

A novel and patented procedure is described for the sonochemical fabrication of a new class of microelectrode array based sensor with electrode element populations of up to 2 x 10(5) cm(-2). For some years it has been accepted that microelectrode arrays offer an attractive route for lowering minimum limits of detection and imparting stir (convectional mass transport) independence to sensor responses; despite this no commercial biosensors, to date, have employed microelectrode arrays, largely due to the cost of conventional fabrication routes that have not proved commercially viable for disposable devices. Biosensors formed by our sonochemical approach offer unrivalled sensitivity and impart stir independence to sensor responses. This format lends itself for mass fabrication due to the simplicity and inexpensiveness of the approach; in the first instance impedimetric and amperometric sensors are reported for glucose as model systems. Sensors already developed for ethanol, oxalate and a number of pesticide determinations will be reported in subsequent publications.     

Beneficial effects of GH/IGF-1 on skeletal muscle atrophy and function in experimental heart failure

Muscle atrophy is a determinant of exercise capacity in heart failure (CHF). Myocyte apoptosis, triggered by tumor necrosis factor- (TNF-) or its second messenger sphingosine (SPH), is one of the causes of atrophy. Growth hormone (GH) improves hemodynamic and cardiac trophism in several experimental models of CHF, but its effect on skeletal muscle in CHF is not yet clear. We tested the hypothesis that GH can prevent skeletal muscle apoptosis in rats with CHF. CHF was induced by injecting monocrotaline. After 2 wk, 2 groups of rats were treated with GH (0.2 mg·kg^–1·day^–1 and 1.0 mg·kg^–1·day^–1) subcutaneously. A third group of controls had saline. After 2 additional weeks, rats were killed. Tibialis anterior cross-sectional area, myosin heavy chain (MHC) composition, and a study on myocyte apoptosis and serum levels of TNF- and SPH were carried out. The number of apoptotic nuclei, muscle atrophy, and serum levels of TNF- and SPH were decreased with GH at high but not at low doses compared with CHF rats. Bcl-2 was increased, whereas activated caspases and bax were decreased. The MHC pattern in GH-treated animals was similar to that of controls. Monocrotaline slowed down both contraction and relaxation but did not affect specific tetanic force, whereas absolute force was decreased. GH treatment restored contraction and relaxation to control values and brought muscle mass and absolute twitch and tetanic tension to normal levels. These findings may provide an insight into the therapeutic strategy of GH given to patients with CHF to improve exercise capacity. apoptosis; cytokines; myosin heavy chains     

Proteomics-based target identification: bengamides as a new class of methionine aminopeptidase inhibitors

LAF389 is a synthetic analogue of bengamides, a class of marine natural products that produce inhibitory effects on tumor growth in vitro and in vivo. A proteomics-based approach has been used to identify signaling pathways affected by bengamides. LAF389 treatment of cells resulted in altered mobility of a subset of proteins on two-dimensional gel electrophoresis. Detailed analysis of one of the proteins, 14-3-3gamma, showed that bengamide treatment resulted in retention of the amino-terminal methionine, suggesting that bengamides directly or indirectly inhibited methionine aminopeptidases (MetAps). Both known MetAps are inhibited by LAF389. Short interfering RNA suppression of MetAp2 also altered amino-terminal processing of 14-3-3gamma. A high resolution structure of human MetAp2 co-crystallized with a bengamide shows that the compound binds in a manner that mimics peptide substrates. Additionally, the structure reveals that three key hydroxyl groups on the inhibitor coordinate the di-cobalt center in the enzyme active site.     

A deficiency screen of the major autosomes identifies a gene (matrimony) that is haplo-insufficient for achiasmate segregation in Drosophila oocytes

In Drosophila oocytes, euchromatic homolog-homolog associations are released at the end of pachytene, while heterochromatic pairings persist until metaphase I. A screen of 123 autosomal deficiencies for dominant effects on achiasmate chromosome segregation has identified a single gene that is haplo-insufficient for homologous achiasmate segregation and whose product may be required for the maintenance of such heterochromatic pairings. Of the deficiencies tested, only one exhibited a strong dominant effect on achiasmate segregation, inducing both X and fourth chromosome nondisjunction in FM7/X females. Five overlapping deficiencies showed a similar dominant effect on achiasmate chromosome disjunction and mapped the haplo-insufficient meiotic gene to a small interval within 66C7-12. A P-element insertion mutation in this interval exhibits a similar dominant effect on achiasmate segregation, inducing both high levels of X and fourth chromosome nondisjunction in FM7/X females and high levels of fourth chromosome nondisjunction in X/X females. The insertion site for this P element lies immediately upstream of CG18543, and germline expression of a UAS-CG18543 cDNA construct driven by nanos-GAL4 fully rescues the dominant meiotic defect. We conclude that CG18543 is the haplo-insufficient gene and have renamed this gene matrimony (mtrm). Cytological studies of prometaphase and metaphase I in mtrm hemizygotes demonstrate that achiasmate chromosomes are not properly positioned with respect to their homolog on the meiotic spindle. One possible, albeit speculative, interpretation of these data is that the presence of only a single copy of mtrm disrupts the function of whatever "glue" holds heterochromatically paired homologs together from the end of pachytene until metaphase I.     

Feeding melatonin enhances the phase shifting response to triazolam in both young and old golden hamsters

Aging alters many aspects of circadian rhythmicity, including responsivity to phase-shifting stimuli and the amplitude of the rhythm of melatonin secretion. As melatonin is both an output from and an input to the circadian clock, we hypothesized that the decreased melatonin levels exhibited by old hamsters may adversely impact the circadian system as a whole. We enhanced the diurnal rhythm of melatonin by feeding melatonin to young and old hamsters. Animals of both age groups on the melatonin diet showed larger phase shifts than control-fed animals in response to an injection with the benzodiazepine triazolam at a circadian time known to induce phase advances in the activity rhythm of young animals. Thus melatonin treatment can increase the sensitivity of the circadian timing system of young animals to a nonphotic stimulus, and the ability to increase this sensitivity persists into old age, indicating exogenous melatonin might be useful in reversing at least some age-related changes in circadian clock function.