Porosomes are the universal secretory portals at the cell plasma membrane, where membrane-bound secretory vesicles transiently dock and fuse to expel intravesicular contents to the outside during cell secretion. In the past decade, the neuronal porosome complex, a 10–15 nm cup-shaped lipoprotein structure has been isolated, its partial composition and 3D contour map determined, and it has been functionally reconstituted into artificial lipid membrane. Here we further determine the composition of the neuronal porosome proteome using immunoisolation and gel filtration chromatography, followed by tandem mass spectrometry. Results from the study demonstrate nearly 40 proteins to constitute the neuronal porosome proteome. Furthermore, interaction of proteins within the porosome and their resulting arrangement is predicted. The association and dissociation of proteins at the porosome following stimulation of cell secretion demonstrates the dynamic nature of the organelle.

Mesenchymal stromal cells (MSCs) have proven useful for cell and immune therapy, but the molecular constituents responsible for their functionalities, in particular, those on the plasma membrane, remain largely unknown. Here we employed both gel and nongel based MS to analyze human MSCs' membrane proteome before and after adipogenesis. 2-DE of cells that were pretreated with membrane impermeable fluorescent dyes revealed that both the whole cell proteome and the cell surface subproteome were independent of donors. LC coupled with tandem MS analysis of the plasma membrane-containing fraction allowed us to identify 707 proteins, approximately half of which could be annotated as membrane-related proteins. Of particular interest was a subset of ectodomain-containing membrane-bound proteins that encompass most known surface markers for MSCs, but also contain a multitude of solute carriers and ATPases. Upon adipogenic differentiation, this proteomic profile was amended to include several proteins involved in lipid metabolism and trafficking, at the expense of, most noticeably, ectoenzymes. Our results here provide not only a basis for future studies of MSC-specific molecular mechanisms, but also a molecular inventory for the development of antibody-based cell isolation and identification procedures.

Immunogold-labeled transmission electron microscopy (TEM) was used to determine the total number of secretory vesicles in resting and in growth hormone (GH)-stimulated porcine pituitary cells. We identified three categories of vesicles: filled, empty, and partly empty. Resting GH cells contained more than twice as many filled vesicles than did the stimulated ones. Stimulated cells, however, contained nearly twice as many empty vesicles and 2.5 times more partly empty vesicles than did resting cells. Secretory vesicles in GH cells further revealed the localization of GH only in electron-dense vesicles in both resting and stimulated cells. The total number of secretory vesicles did not change after secretion. These results are consistent with a mechanism that, after stimulation of secretion, vesicles transiently dock and fuse at the fusion pore to release vesicular contents.

BackgroundThere is growing evidence that inflammatory processes of activated microglia could play an important role in the progression of nerve cell damage in neurodegenerative disorders such as Parkinson’s disease and Alzheimer’s disease which harbor features of chronic microglial activation, though the precise mechanism is unknown. In this study, we presented in vivo and ex vivo experimental evidences indicating that activated microglia could exacerbate the survival of axotomized dopaminergic neurons and that appropriate inactivation of microglia could be neuroprotective.ResultsThe transection of medial forebrain bundle (MFB) of a rat induced loss of dopaminergic neurons in a time-dependent manner and accompanied with microglial activation. Along with microglial activation, production of reactive oxygen species (ROS) was upregulated and TH/OX6/hydroethidine triple-immunofluorescence showed that the microglia mainly produced ROS. When the activated microglial cells that were isolated from the substantia nigra of the MFB axotomized animal, were transplanted into the substantia nigra of which MFB had been transected at 7 days ago, the survival rate of axotomized dopaminergic neurons was significantly reduced as compared with sham control. Meanwhile, when the microglial activation was attenuated by administration of tuftsin fragment 1-3 (microglia inhibitory factor) into the lateral ventricle using mini-osmotic pump, the survival rate of axotomized dopaminergic neurons was increased.ConclusionThe present study suggests that activated microglia could actively produce and secrete unfavorable toxic substances, such as ROS, which could accelerate dopaminergic neuronal cell loss. So, well-controlled blockade of microglial activation might be neuroprotective in some neuropathological conditions.

This study aimed to systematically analyse the differences in emotions evoked by the same colour, depending on whether it is an object or light colour. To this end, an assessment was conducted using mock experimental spaces created with either object or light colours (red, green, blue and yellow) and with different levels of illuminance (10 lx or 100 lx). The participants comprised 30 graduate students (19 men; 20–30 years) majoring in colour and lighting, and the vocabulary selected for emotional assessment included the words ‘comfortable’, ‘stable’, ‘tense’, ‘anxious’, ‘cool’ and ‘warm’. The results demonstrated differences between participants’ emotional reactions to the object colour and light colour environments when the same colour was used. For red, green and blue, the object colour was rated higher than the light colour for positive responses of ‘comfortable’ and ‘stable’. For yellow, however, the response differed only slightly between object colour and light colour. Finally, emotional differences varied significantly depending on illuminance level for object colour but not for light colour.

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