Aggregatibacter actinomycetemcomitans induces detachment and death of human gingival epithelial cells and fibroblasts via elastase release following leukotoxin‐dependent neutrophil lysis

Aggregatibacter actinomycetemcomitans is considered to be associated with periodontitis. Leukotoxin (LtxA), which destroys leukocytes in humans, is one of this bacterium's major virulence factors. Amounts of neutrophil elastase (NE), which is normally localized in the cytoplasm of neutrophils, are reportedly increased in the saliva of patients with periodontitis. However, the mechanism by which NE is released from human neutrophils and the role of NE in periodontitis is unclear. In the present study, it was hypothesized that LtxA induces NE release from human neutrophils, which subsequently causes the breakdown of periodontal tissues. LtxA‐treatment did not induce significant cytotoxicity against human gingival epithelial cells (HGECs) or human gingival fibroblasts (HGFs). However, it did induce significant cytotoxicity against human neutrophils, leading to NE release. Furthermore, NE and the supernatant from LtxA‐treated human neutrophils induced detachment and death of HGECs and HGFs, these effects being inhibited by administration of an NE inhibitor, sivelestat. The present results suggest that LtxA mediates human neutrophil lysis and induces the subsequent release of NE, which eventually results in detachment and death of HGECs and HGFs. Thus, LtxA‐induced release of NE could cause breakdown of periodontal tissue and thereby exacerbate periodontitis.     

Age-related differences in muscle activity, stride frequency and heart rate response during walking in water.

This study aimed to examine whether walking in water produces age-related differences in muscle activity, stride frequency (SF), and heart rate (HR) response. Surface electromyography (EMG) was used to evaluate muscle activities in six older and six young subjects while they walked in water immersed to the level of the xiphoid process. The trials in water utilized the Flowmill which consists of a treadmill at the base of a water flume. The measurement of maximal voluntary contraction (MVC) of each muscle was made prior to the gait analysis. The %MVCs, which refer to the surface EMG measures, from the gastrocnemius of the older subjects were significantly lower than those of the young subjects, in every experimental condition (P<0.05). In contrast, the %MVCs from the rectus femoris (P<0.05) and the biceps femoris (P<0.001) of older subjects were significantly greater than those of young subjects in every experimental condition. Moreover, the SFs of older subjects were also significantly greater than those of young subjects (P<0.05), while the HR responses of older and young subjects were similar. In conclusion, the older subjects had increased hip musculature activity and decreased ankle plantar flexor activity while walking in water, compared with the young subjects.     

Localization of spectrin isoforms in the adult mouse heart

The distribution of two isoforms of spectrin in the adult mouse heart was investigated by Western blotting and immunocytochemistry by use of monospecific antibodies to erythrocyte spectrin and nonerythroid brain spectrin (240/235). Western blotting revealed proteins analogous to both isoforms of -spectrin in adult heart. Light-microscopic immunocytochemistry indicated that erythroid spectrin was distributed throughout the myocardium, with immunofluorescence localized to plasma membranes, Z-lines, and intercalated discs. Antibodies to brain spectrin (240/235) exhibited staining throughout the heart, with a generally diffuse distribution except for the prominent immunoreactivity associated with the intercalated discs. Nonerythroid spectrin immunofluorescence was detected in the endothelial cells of the endocardium and the mesothelial cell lining of the epicardium. Erythrocyte spectrin was not detected in the endocardium or the epicardium. The identification and localization of spectrin isoforms in the mammalian heart suggest the importance of spectrin proteins in the structural integrity and proper function of cardiac cells and tissues. This is the first demonstration of two different -spectrin subunits in the mammalian heart.     

A Simple Method for Making a Vibrating Probe System

A simple and easy method for making the vibrating probe systemfirst developed by Jaffe and Nuccitelli [(1974) J. Cell Biol.63: 614] is described. The best performance of the probe insea water was 30.7 µA cm^-2 per one µV output, 0.4µA cm^-2 peak-to-peak output-noise and negligible output-driftat 30-µm vibrating amplitude and 3-s amplifier time constant.Sample applications were made for both freshwater and marinealgae. (Received March 22, 1984; Accepted May 28, 1984)     

Ionic composition and pH of the vacuolar sap in marine dinoflagellate Noctiluca

Ionic composition of the vacuolar sap of Noctiluca miliariswas as follows: [Na⁺] = 487.3 mM, [K⁺]=24.1 mM, [Ca²⁺]=6.6 mM,[Mg²⁺]=2.8 mM, [Cl^–]=500mM, [NH₄⁺]=15–25 mM, and[SO₄^2–]=undetectable. To measure the vacuolar pH of singleliving cells, a pH-sensitive glass microelectrode was used.The vacuolar pH value was 3.50 ±0.18. When the cellswere transferred from normal sea water into osmotically adjusted50% sea water for one day, the vacuolar ion concentrations remainedalmost constant. Upon immersing the cells in osmotically unadjustedsea water of various concentrations for one day, the observedincrements or decrements of the vacuolar ion concentrationscould be accounted for largely by the migration of water outof or into the cells. The intrinsic ionic composition of thevacuole seems to be constant against changes in ion concentrationsof the bathing medium. (Received October 20, 1975; )     

Neural circuit mechanisms of hierarchical sequence learning tested on large-scale recording data

The brain performs various cognitive functions by learning the spatiotemporal salient features of the environment. This learning requires unsupervised segmentation of hierarchically organized spike sequences, but the underlying neural mechanism is only poorly understood. Here, we show that a recurrent gated network of neurons with dendrites can efficiently solve difficult segmentation tasks. In this model, multiplicative recurrent connections learn a context-dependent gating of dendro-somatic information transfers to minimize error in the prediction of somatic responses by the dendrites. Consequently, these connections filter the redundant input features represented by the dendrites but unnecessary in the given context. The model was tested on both synthetic and real neural data. In particular, the model was successful for segmenting multiple cell assemblies repeating in large-scale calcium imaging data containing thousands of cortical neurons. Our results suggest that recurrent gating of dendro-somatic signal transfers is crucial for cortical learning of context-dependent segmentation tasks.     

Aberrant Active cis-Regulatory Elements Associated with Downregulation of RET Finger Protein Overcome Chemoresistance in Glioblastoma

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The inositol 5-phosphatase SHIP2 is an effector of RhoA and is involved in cell polarity and migration

Cell migration is essential for various physiological and pathological processes. Polarization in motile cells requires the coordination of several key signaling molecules, including RhoA small GTPases and phosphoinositides. Although RhoA participates in a front-rear polarization in migrating cells, little is known about the functional interaction between RhoA and lipid turnover. We find here that src-homology 2-containing inositol-5-phosphatase 2 (SHIP2) interacts with RhoA in a GTP-dependent manner. The association between SHIP2 and RhoA is observed in spreading and migrating U251 glioma cells. The depletion of SHIP2 attenuates cell polarization and migration, which is rescued by wild-type SHIP2 but not by a mutant defective in RhoA binding. In addition, the depletion of SHIP2 impairs the proper localization of phosphatidylinositol 3,4,5-trisphosphate, which is not restored by a mutant defective in RhoA binding. These results suggest that RhoA associates with SHIP2 to regulate cell polarization and migration.     

REFOLDdb: a new and sustainable gateway to experimental protocols for protein refolding

Background

More than 7000 papers related to “protein refolding” have been published to date, with approximately 300 reports each year during the last decade. Whilst some of these papers provide experimental protocols for protein refolding, a survey in the structural life science communities showed a necessity for a comprehensive database for refolding techniques. We therefore have developed a new resource – “REFOLDdb” that collects refolding techniques into a single, searchable repository to help researchers develop refolding protocols for proteins of interest.

Results

We based our resource on the existing REFOLD database, which has not been updated since 2009. We redesigned the data format to be more concise, allowing consistent representations among data entries compared with the original REFOLD database. The remodeled data architecture enhances the search efficiency and improves the sustainability of the database. After an exhaustive literature search we added experimental refolding protocols from reports published 2009 to early 2017. In addition to this new data, we fully converted and integrated existing REFOLD data into our new resource. REFOLDdb contains 1877 entries as of March 17^th, 2017, and is freely available at http://p4d-info.nig.ac.jp/refolddb/.

Conclusion

REFOLDdb is a unique database for the life sciences research community, providing annotated information for designing new refolding protocols and customizing existing methodologies. We envisage that this resource will find wide utility across broad disciplines that rely on the production of pure, active, recombinant proteins. Furthermore, the database also provides a useful overview of the recent trends and statistics in refolding technology development.

     

Autoregulation of Osteocyte Sema3A Orchestrates Estrogen Action and Counteracts Bone Aging

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