Sphingosylphosphorylcholine generates reactive oxygen species through calcium-, protein kinase Cdelta- and phospholipase D-dependent pathways

Sphingosylphosphorylcholine (SPC) is a bioactive lipid molecule involved in numerous biological processes. Treatment of MS1 pancreatic islet endothelial cells with SPC increased phospholipase D (PLD) activity in a time- and dose-dependent manner. In addition, treatment of the MS1 cells with 10 microM SPC induced stimulation of phospholipase C (PLC) activity and transient elevation of intracellular Ca2+. The SPC-induced PLD activation was prevented by pretreatment of the MS1 cells with a PLC inhibitor, U73122, and an intracellular Ca2+-chelating agent, BAPTA-AM. This suggests that PLC-dependent elevation of intracellular Ca2+ is involved in the SPC-induced activation of PLD. The SPC-dependent PLD activity was also almost completely prevented by pretreatment with pan-specific PKC inhibitors, GF109203X and RO-31-8220, and with a PKCdelta-specific inhibitor, rottlerin, but not by pretreatment with GO6976, a conventional PKC isozymes-specific inhibitor. Adenoviral overexpression of a kinase-deficient mutant of PKCdelta attenuated the SPC-induced PLD activity. These results suggest that PKCdelta plays a crucial role for the SPC-induced PLD activation. The SPC-induced PLD activation was preferentially potentiated in COS-7 cells transfected with PLD2 but not with PLD1, suggesting a specific implication of PLD2 in the SPC-induced PLD activation. SPC treatment induced phosphorylation of PLD2 in COS-7 cells, and overexpression of the kinase-deficient mutant of PKCdelta prevented the SPC-induced phosphorylation of PLD2. Furthermore, SPC treatment generated reactive oxygen species (ROS) in MS1 cells and the SPC induced production of ROS was inhibited by pretreatment with U73122, BAPTA-AM, and rottlerin. In addition, pretreatment with a PLD inhibitor 1-butanol and overexpression of a lipase-inactive mutant of PLD2 but not PLD1 attenuated the SPC-induced generation of ROS. These results suggest that PLC-, Ca2+-, PKCdelta-, and PLD2-dependent pathways are essentially required for the SPC induced ROS generation.     

Differential effects of circadian typology on sleep-related symptoms,physical fatigue and psychological well-being in relation to resilience

Various physiological and psychological functions are influenced by circadian typology (CT), which was reported to be related to resilience. However, few studies have assessed the effects of CT in relation to resilience. The aim of the present study was to assess the influence of CT on sleep-related symptoms, physical fatigue and psychological well-being in relation to resilience. The present study included a total of 1794 healthy hospital employees, and they completed the Morningness–Eveningness Questionnaire, Connor–Davidson Resilience Scale, Pittsburgh Sleep Quality Index, Epworth Sleepiness Scale, Fatigue Severity Scale, Hospital Anxiety and Depression Scale and World Health Organization Quality of Life Scale Abbreviated Version. Subjects with evening type showed lower sleep quality, more daytime sleepiness and physical fatigue than neither types and morning types. Additionally, evening types were more depressed and anxious and reported a poorer quality of life. CT was found to be a significant predictor of sleep quality, but CT was minimally associated with physical fatigue and psychological well-being in the regression analysis. Instead, resilience was substantially related to all of the variables measured. In conclusion, CT independently predicts sleep quality, but the effects of CT on physical fatigue and psychological well-being are negligible compared to those of resilience.     

Eclalbasaponin II Ameliorates the Cognitive Impairment Induced by Cholinergic Blockade in Mice

Eclalbasaponin II derived from Eclipta prostrata L. (Asteraceae) has been reported to have anti-fibrotic, anti-bacterial and autophagic activities, but its effect on cognitive function has not been investigated. We studied the effect of eclalbasaponin II on cholinergic blockade-induced memory impairment in mice using the passive avoidance, Y-maze, and Morris water maze tasks. Eclalbasaponin II (10 or 20 mg/kg, p.o.) significantly ameliorated the cognitive dysfunction induced by scopolamine in the passive avoidance, Y-maze, and the Morris water maze tasks. To identify the mechanism of the memory-ameliorating effect of eclalbasaponin II, acetylcholinesterase (AChE) activity assay, Western blot analysis and electrophysiology were conducted. Eclalbasaponin II inhibited the AChE activity in ex vivo study, and the administration of eclalbasaponin II and its metabolite, echinocystic acid, increased the phosphorylation levels of memory-related signaling molecules, including protein kinase B (Akt) and glycogen synthase kinase-3β (GSK-3β), in the hippocampus. Although eclalbasaponin II did not affect hippocampal long term potentiation (LTP), echinocystic acid significantly enhanced hippocampal LTP formation (30 μM). These results suggest that eclalbasaponin II ameliorates cholinergic blockade-induced cognitive impairment via AChE inhibition, LTP formation and the activation of Akt-GSK-3β signaling, and that eclalbasaponin II may be a useful to treat cognitive impairment derived from cholinergic dysfunction.     

Rhodeus pseudosericeus sp. nov., a new bitterling from South Korea (Cyprinidae, Acheilognathinae)

A new bitterling, Rhodeus pseudosericeus sp. nov., is described on the basis of 31 specimens from five localities included in the Namhan River system, South Korea. The new species is distinguished from other Rhodeus species by the following combination of characters: branched dorsal fin rays 9–10 (mode 9); branched anal fin rays 9–11 (mode 10); longest simple ray of dorsal fin strong and stiff, distally segmented; pelvic fin rays i, 6–7; iris of males blackish; dorsal and anal fins of males grayish in breeding season; karyotype with 2n = 48 (8m + 20sm + 20st). Rhodeus pseudosericeus sp. nov. is similar to Rhodeus sericeus sericeus in the number of pelvic fin and branched dorsal fin rays and the melanophores present on the dorsal fin membrane, but differs from the latter in having a greater body depth, more branched anal fin rays, fewer vertebrae, a lower number of scales in the lateral series, and differing male nuptial coloration. Received: June 30, 2000 / Revised: February 21, 2001 / Accepted: March 6, 2001     

Expression of gibberellin 2-oxidase 4 from Arabidopsis under the control of a senescence-associated promoter results in a dominant semi-dwarf plant with normal flowering

Gibberellin (GA), a plant hormone, is involved in many aspects of plant growth and development both in vegetative and reproductive phases. GA2-oxidase plays a key role in the GA catabolic pathway to reduce bioactive GAs. We produced transgenic Arabidopsis plants expressing GA2-oxidase 4 (AtGA2ox4) under the control of a senescenceassociated promoter (SEN1). As we hypothesized, transgenic plants (SEN1::AtGA2ox4) exhibited a dominant semi-dwarf phenotype with a decrease of bioactive GAs (e.g., GA4 and GA1) up to two-fold compared to control plants. Application of bioactive GA3 resulted in increased shoot length, indicating that the GA signaling pathway functions normally in the SEN1::AtGA2ox4 plants. Expressions of other members of GA2-oxidase family, such as AtGA2ox1, AtGA2ox3, AtGA2ox6, and AtGA2ox8, were decreased slightly in the flower and silique tissues while GA biosynthetic genes (e.g., AtGA20ox1, AtGA20ox2 and AtGA3ox1) were not significantly changed in the SEN::AtGA2ox4 plants. Using proteome profiling (2-D PAGE followed by MALDI-TOF/MS), we identified 29 protein spots that were increased in the SEN1::AtGA2ox4 plants, but were decreased to wild-type levels by GA3 treatment. The majority were found to be involved in photosynthesis and carbon/energy metabolism. Unlike the previous constitutive over-expression of GA2-oxidases, which frequently led to floral deformity and/or loss of fertility, the SEN1::AtGA2ox4 plants retained normal floral morphology and seed production. Accordingly, the expressions of FT and CO genes remained unchanged in the SEN1::AtGA2ox4 plants. Taken together, our results suggest that the dominant dwarf trait carried by SEN1::AtGA2ox4 plants can be used as an efficient dwarfing tool in plant biotechnological applications.     

Phosphorylation of focal adhesion kinase at tyrosine 861 is crucial for Ras transformation of fibroblasts

Although elevated expression and increased tyrosine phosphorylation of focal adhesion kinase (FAK) are crucial for tumor progression, the mechanism by which FAK promotes oncogenic transformation is unclear. We have therefore determined the role of FAK phosphorylation at tyrosine 861 in the oncogenic transformation of NIH3T3 fibroblasts. FAK phosphorylation at tyrosine 861 was increased in both constitutively H-Ras-transformed and H-Ras-inducible NIH3T3 cells, in parallel with cell transformation. However, H-Ras-inducible cells transfected with the nonphosphorylatable mutant FAK Y861F showed decreased migration/invasion, focus forming activity and anchorage-independent growth, compared with either wild-type or kinase-defective FAK. In contrast to unaltered FAK/Src activity, the association of FAK and p130(CAS) was decreased in FAK Y861F-transfected cells, and FAK phosphorylation at tyrosine 861 enhanced this association in vitro. Consistently, FAK Y861F-transfected cells were defective in activation of c-Jun NH(2)-terminal kinase and in expression of matrix metalloproteinase-9 during transformation. Taken together, these results strongly suggest that FAK phosphorylation at tyrosine 861 is crucial for H-Ras-induced transformation through regulation of the association of FAK with p130(CAS).     

Effect of JGK-263 as a new glycogen synthase kinase-3β inhibitor on extrinsic apoptosis pathway in motor neuronal cells

Glycogen synthase kinase-3β (GSK-3β) has been identified as one of the important pathogenic mechanisms in motor neuronal death. GSK-3β inhibitor has been investigated as a modulator of apoptosis and has been shown to confer significant protective effects on cell death in neurodegenerative diseases. However, GSK-3β is known to have paradoxical effects on apoptosis subtypes, i.e., pro-apoptotic in mitochondrial-associated intrinsic apoptosis, but anti-apoptotic in death receptor-related extrinsic apoptosis. In this study, we evaluated the effect of a new GSK-3β inhibitor (JGK-263) on motor neuron cell survival and apoptosis, by using low to high doses of JGK-263 after 48 h of serum withdrawal, and monitoring changes in extrinsic apoptosis pathway components, including Fas, FasL, cleaved caspase-8, p38α, and the Fas–Daxx interaction. Cell survival peaked after treatment of serum-deprived cells with 50 μM JGK-263. The present study showed that treatment with JGK-263 reduced serum-deprivation-induced motor neuronal apoptosis by inactivating not only the intrinsic, but also the extrinsic apoptosis pathway. These results suggest that JGK-263 has a neuroprotective effect through effective modulation of the extrinsic apoptosis pathway in motor neuron degeneration.