2-DE profiling of GDNF overexpression-related proteome changes in differentiating ST14A rat progenitor cells

Targeted differentiation of neural progenitor cells (NPCs) is a challenge for treatment of neurodegenerative diseases by cell replacement therapy and cell signalling manipulation. Here, we applied a proteome profiling approach to the rat striatal progenitor model cell line ST14A in order to elucidate cellular differentiation processes. Native cells and cells transfected with the glial cell line-derived neurotrophic factor (GDNF) gene were investigated at the proliferative state and at seven time points up to 72 h after induction of differentiation. 2-DE combined with MALDI-MS was used to create a reference 2-DE-map of 652 spots of which 164 were identified and assigned to 155 unique proteins. For identification of protein expression changes during cell differentiation, spot patterns of triplicate gels were matched to the 2-DE-map. Besides proteins that display expression changes in native cells, we also noted 43 protein-spots that were differentially regulated by GDNF overexpression in more than four time points of the experiment. The expression patterns of putative differentiation markers such as annexin 5 (ANXA5), glucosidase II beta subunit (GLU2B), phosphatidylethanolamine-binding protein 1 (PEBP1), myosin regulatory light chain 2-A (MLRA), NASCENT polypeptide-associated complex alpha (NACA), elongation factor 2 (EF2), peroxiredoxin-1 (PRDX1) and proliferating cell nuclear antigen (PCNA) were verified by Western blotting. The results reflect the large rearrangements of the proteome during the differentiation process of NPCs and their strong modification by neurotrophic factors like GDNF.     

Sleep, sleepiness, fatigue, and performance of 12-hour-shift nurses

Nurses working 12-h shifts complain of fatigue and insufficient/poor-quality sleep. Objectively measured sleep times have not been often reported. This study describes sleep, sleepiness, fatigue, and neurobehavioral performance over three consecutive 12-h (day and night) shifts for hospital registered nurses. Sleep (actigraphy), sleepiness (Karolinska Sleepiness Scale [KSS]), and vigilance (Performance Vigilance Task [PVT]), were measured serially in 80 registered nurses (RNs). Occupational fatigue (Occupational Fatigue Exhaustion Recovery Scale [OFER]) was assessed at baseline. Sleep was short (mean 5.5?h) between shifts, with little difference between day shift (5.7?h) and night shift (5.4?h). Sleepiness scores were low overall (3 on a 1-9 scale, with higher score indicating greater sleepiness), with 45% of nurses having high level of sleepiness (score >?7) on at least one shift. Nurses were progressively sleepier each shift, and night nurses were sleepier toward the end of the shift compared to the beginning. There was extensive caffeine use, presumably to preserve or improve alertness. Fatigue was high in one-third of nurses, with intershift fatigue (not feeling recovered from previous shift at the start of the next shift) being most prominent. There were no statistically significant differences in mean reaction time between day/night shift, consecutive work shift, and time into shift. Lapsing was traitlike, with rare (39% of sample), moderate (53%), and frequent (8%) lapsers. Nurses accrue a considerable sleep debt while working successive 12-h shifts with accompanying fatigue and sleepiness. Certain nurses appear more vulnerable to sleep loss than others, as measured by attention lapses.     

Effects of the juvenile hormone analog pyriproxyfen on German cockroach, Blattella germanica (L.), tergal gland development and production of tergal gland secretion proteins

Male German cockroaches possess secretory glands that secrete fluid into a pair of transverse depressions on the seventh and eighth abdominal tergites. We investigated the effects of altered juvenoid titer during the first part of the last instar on tergal gland secretory tissue development and the production of tergal gland secretion proteins. Male fifth (last) instar nymphs (1-3 days post-emergent) were topically treated with the JH analog pyriproxyfen. Light and transmission electron microscopy demonstrated that treatment with pyriproxyfen produced a visible decrease in the amount of tergal gland tissue present, a deformation of the overall shape of the gland located on tergite seven, and a less orderly arrangement of the secretory cells in the tissue. The protein fraction of tergal gland secretion was examined in pyriproxyfen-treated and control insects 1, 5, and 15 days after the insects molted to the adult stage. Amounts of all tergal secretion proteins were reduced in treated insects.     

Consequences of food distribution for optimal searching behavior: an evolutionary model

Resource distribution can vary greatly in space and time. Consequently, animals should adjust their searching tactics to such spatio–temporal patterns in accordance with their innate capabilities, or alternatively, they should use a genetically fixed searching tactic that has been evolved in response to the specific pattern of the food they experience. Using a simulation model and a genetic algorithm, we show how optimal searching tactics change as a function of food spatial pattern. Searching tactics for hidden prey can be approximated using the following three components: (1) Extensive search mode (ESM), the type of movement before encountering a food item; (2) Intensive search mode (ISM), the type of movement after encountering a food item; and (3) ISM duration. Both ESM and ISM are characterized by movement tortuosity. We show that searching behavior adaptively changes as a function of food pattern. When food is distributed in a regular pattern, ISM is more directional than ESM, but under a clumped food pattern, ISM is much more tortuous than ESM. It may suggest that animals with larger spectra of searching tactics should experience greater variance or seasonal changes in their food pattern than animals with narrow spectra of searching tactics. Increased forager attack radius diminishes the differences between ESM and ISM, and thus the use of these three components to model searching in animals with higher attack radii is not appropriate. Increased handling time, which is a surrogate of reducing habitat profitability results in longer patch residency time as expected by optimal foraging theory. To conclude, we suggest that using such a combined approach of simulation models and genetic algorithms may improve our understanding of how extrinsic and intrinsic factors interact to influence searching behavior.     

Dose-dependent effect of radiation on angiogenic and angiostatic CXC chemokine expression in human endothelial cells

Blood vessel growth is regulated by angiogenic and angiostatic CXC chemokines, and radiation is a vasculogenic stimulus. We investigated the effect of radiation on endothelial cell chemokine signaling, receptor expression, and migration and apoptosis. Human umbilical vein endothelial cells were exposed to a single fraction of 0, 5, or 20 Gy of ionizing radiation (IR). All vasculogenic chemokines (CXCL1–3/5–8) increased 3–13-fold after 5 or 20 Gy IR. 20 Gy induced a marked increase (1.6–4-fold) in angiostatic CXC chemokines. CXCR4 expression increased 3.5 and 7-fold at 48 h after 5 and 20 Gy, respectively. Bone marrow progenitor cell chemotaxis was augmented by conditioned media from cells treated with 5 Gy IR. Whereas 5 Gy markedly decreased intrinsic cell apoptosis (0 Gy = 16% ± 3.6 vs. 5 Gy = 4.5% ± 0.3), 20 Gy increased it (21.4% ± 1.2); a reflection of pro-survival angiogenic chemokine expression. Radiation induces a dose-dependent increase in pro-angiogenic CXC chemokines and CXCR4. In contrast, angiostatic chemokines and apoptosis were induced at higher (20 Gy) radiation doses. Cell migration improved significantly following 5 Gy, but not 20 Gy IR. Collectively, these data suggest that lower doses of IR induce an angiogenic cascade while higher doses produce an angiostatic profile.     

Production and characterization of a recombinant beta‐1,4‐endoglucanase (glycohydrolase family 9) from the termite Reticulitermes flavipes

Cell‐1 is a host‐derived beta‐1,4‐endoglucanase (Glycohydrolase Family 9 [GHF9]) from the lower termite Reticulitermes flavipes. Here, we report on the heterologous production of Cell‐1 using eukaryotic (Baculovirus Expression Vector System; BEVS) and prokaryotic (E. coli) expression systems. The BEVS‐expressed enzyme was more readily obtained in solubilized form and more active than the E. coli–expressed enzyme. K_m and V_max values for BEVS‐expressed Cell‐1 against the model substrate CMC were 0.993% w/v and 1.056 µmol/min/mg. Additional characterization studies on the BEVS‐expressed enzyme revealed that it possesses activity comparable to the native enzyme, is optimally active around pH 6.5–7.5 and 50–60°C, is inhibited by EDTA, and displays enhanced activity up to 70°C in the presence of CaCl₂. These findings provide a foundation on which to begin subsequent investigations of collaborative digestion by coevolved host and symbiont digestive enzymes from R. flavipes that include GHF7 exoglucanases, GHF1 beta glucosidases, phenol‐oxidizing laccases, and others. © 2010 Wiley Periodicals, Inc.