• MeSH
• chloramfenikol-O-acetyltransferasa MeSH
• chromatografie na tenké vrstvě MeSH
• genetická transkripce MeSH
• geny MHC třídy I MeSH
• myši MeSH
• regulace genové exprese MeSH
• reportérové geny MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• přehledy MeSH

• MeSH
• exprese genu MeSH
• tranzitorní ischemická ataka genetika   MeSH

The transient receptor potential ion channel TRPA1 is a Ca2+-permeable nonselective cation channel widely expressed in sensory neurons, but also in many nonneuronal tissues typically possessing barrier functions, such as the skin, joint synoviocytes, cornea, and the respiratory and intestinal tracts. Here, the primary role of TRPA1 is to detect potential danger stimuli that may threaten the tissue homeostasis and the health of the organism. The ability to directly recognize signals of different modalities, including chemical irritants, extreme temperatures, or osmotic changes resides in the characteristic properties of the ion channel protein complex. Recent advances in cryo-electron microscopy have provided an important framework for understanding the molecular basis of TRPA1 function and have suggested novel directions in the search for its pharmacological regulation. This chapter summarizes the current knowledge of human TRPA1 from a structural and functional perspective and discusses the complex allosteric mechanisms of activation and modulation that play important roles under physiological or pathophysiological conditions. In this context, major challenges for future research on TRPA1 are outlined.

• MeSH
• alosterická regulace MeSH
• elektronová kryomikroskopie metody   MeSH
• kationtové kanály TRP metabolismus   chemie   fyziologie   MeSH
• kationtový kanál TRPA1 * metabolismus   chemie   fyziologie   MeSH
• lidé MeSH
• vztahy mezi strukturou a aktivitou MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

• MeSH
• finanční podpora výzkumu jako téma MeSH
• Gaucherova nemoc enzymologie   MeSH
• genetické vektory MeSH
• glukosylceramidasa biosyntéza   genetika   MeSH
• klonování DNA MeSH
• lidé MeSH
• mutace MeSH
• virus vakcinie MeSH
• Check Tag
• lidé MeSH

Anionic phospholipids represent only minor fraction of cell membranes lipids but they are critically important for many membrane-related processes, including membrane identity, charge, shape, the generation of second messengers, and the recruitment of peripheral proteins. The main anionic phospholipids of the plasma membrane are phosphoinositides phosphatidylinositol 4-phosphate (PI4P), phosphatidylinositol 4,5-bisphosphate (PI4,5P2), phosphatidylserine (PS), and phosphatidic acid (PA). Recent insights in the understanding of the nature of protein-phospholipid interactions enabled the design of genetically encoded fluorescent molecular probes that can interact with various phospholipids in a specific manner allowing their imaging in live cells. Here, we describe the use of transiently transformed plant cells to study phospholipid-dependent membrane recruitment.

• MeSH
• exprese genu MeSH
• fluorescenční barviva analýza   metabolismus   MeSH
• fluorescenční mikroskopie metody   MeSH
• fosfatidylinositoly analýza   metabolismus   MeSH
• fosfolipidy analýza   metabolismus   MeSH
• konfokální mikroskopie metody   MeSH
• luminescentní proteiny analýza   genetika   MeSH
• pyl chemie   genetika   MeSH
• rostlinné buňky chemie   metabolismus   MeSH
• tabák chemie   cytologie   genetika   MeSH
• transformace genetická MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Ulinastatin [or called as urinary trypsin inhibitor (UTI)] plays a role in regulating neurological deficits evoked by transient cerebral ischemia. However, the underlying mechanisms still need to be determined. The present study was to examine the effects of UTI on autophagy, Nrf2-ARE and apoptosis signal pathway in the hippocampus in the process of neurological functions after cerebral ischemia using a rat model of cardiac arrest (CA). CA was induced by asphyxia followed by cardiopulmonary resuscitation (CPR) in rats. Western blot analysis was employed to determine the expression of representative autophagy (namely, Atg5, LC3, Beclin 1), p62 protein (a maker of autophagic flux), and Nrf2-ARE pathways. Neuronal apoptosis was assessed by determining expression levels of Caspase-3 and Caspase-9, and by examining terminal deoxynucleotide transferase-mediated dUTP nick-end labeling (TUNEL). The modified neurological severity score (mNSS) and spatial working memory performance were used to assess neurological deficiencies in CA rats. Our results show that CA amplified autophagy and apoptotic Caspase-3/Caspase-9, and downregulated Nrf2-ARE pathway in the hippocampus CA1 region. Systemic administration of UTI attenuated autophagy and apoptosis, and largely restored Nrf2-ARE signal pathway following cerebral ischemia and thereby alleviated neurological deficits with increasing survival of CA rats. Our data suggest that UTI improves the worsened protein expression of autophagy and apoptosis, and restores Nrf2-ARE signals in the hippocampus and this is linked to inhibition of neurological deficiencies in transient cerebral ischemia. UTI plays a beneficial role in modulating neurological deficits induced by transient cerebral ischemia via central autophagy, apoptosis and Nrf2-ARE mechanisms.

• MeSH
• antioxidační responzivní elementy účinky léků   fyziologie   MeSH
• apoptóza účinky léků   fyziologie   MeSH
• autofagie účinky léků   fyziologie   MeSH
• faktor 2 související s NF-E2 metabolismus   MeSH
• glykoproteiny farmakologie   terapeutické užití   MeSH
• hipokampus účinky léků   metabolismus   patologie   MeSH
• inhibitory trypsinu farmakologie   terapeutické užití   MeSH
• krysa rodu rattus MeSH
• náhodné rozdělení MeSH
• neuroprotektivní látky farmakologie   terapeutické užití   MeSH
• potkani Sprague-Dawley MeSH
• signální transdukce účinky léků   fyziologie   MeSH
• tranzitorní ischemická ataka farmakoterapie   metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Vagal afferents regulate numerous physiological functions including arterial blood pressure, heart rate, breathing, and nociception. Cell bodies of vagal afferents reside in the inferior vagal (nodose) ganglia and their stimulation by various means is being considered as a way to regulate cardiorespiratory responses and control pain sensations. Stimulation of the nodose by exposure to infrared light is recently being considered as a precise way to elicit responses. These responses would likely involve the activity of temperature-sensitive membrane-bound channels. While papers have been published to track the expression of these transient receptor potential ion channels (TRPs), further studies are warranted to determine the in situ expression of the endogenous TRP proteins in the nodose ganglia to fully understand their pattern of expression, subcellular locations, and functions in this animal model. TRP ion channels are a superfamily of Na+ /Ca2+ -channels whose members are temperature- and/or mechano-sensitive and therefore represent a potential set of proteins that will be activated directly or indirectly by infrared light. Here, we report the spatial localization of six TRP channels, TRPV1, TRPV4, TRPM3, TRPM8, TRPA1, and TRPC1, from nodose ganglia taken from juvenile male Sprague-Dawley rats. The channels were detected using immunohistology with fluorescent tags on cryosections and imaged using confocal microscopy. All six TRP channels were detected with different levels of intensity in neuronal cell bodies and some were also detected in axonal fibers and blood vessels. The TRP receptors differed in their prevalence, in their patterns of expression, and in subcellular expression/localization. More specifically, TRPV1, TRPV4, TRPA1, TRPM8, TRPC1, and TRPM3 were found in vagal afferent cell bodies with a wide range of immunostaining intensity from neuron to neuron. Immunostaining for TRPV1, TRPV4, and TRPA1 appeared as fine particles scattered throughout the cytoplasm of the cell body. Intense TRPV1 immunostaining was also evident in a subset of axonal fibers. TRPM8 and TRPC1 were expressed in courser particles suggesting different subcellular compartments than for TRPV1. The localization of TRPM3 differed markedly from the other TRP channels with an immunostaining pattern that was localized to the periphery of a subset of cell bodies, whereas a scattering or no immunostaining was detected within the bulk of the cytoplasm. TRPV4 and TRPC1 were also expressed on the walls of blood vessels. The finding that all six TRP channels (representing four subfamilies) were present in the nodose ganglia provides the basis for studies designed to understand the roles of these channels in sensory transmission within vagal afferent fibers and in the responses elicited by exposure of nodose ganglia to infrared light and other stimuli. Depending on the location and functionality of the TRP channels, they may regulate the flux of Na+ /Ca2+ -across the membranes of cell bodies and axons of sensory afferents, efferent (motor) fibers coursing through the ganglia, and in vascular smooth muscle.

• MeSH
• ganglion inferius metabolismus   MeSH
• kationtové kanály TRP * metabolismus   MeSH
• kationtové kanály TRPM * metabolismus   MeSH
• kationtové kanály TRPV MeSH
• krysa rodu rattus MeSH
• nervus vagus metabolismus   MeSH
• potkani Sprague-Dawley MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Keratin 19 and nuclear reactivity to an endogenous lectin, galectin-1, represent a potential marker of epidermal stem cells. We detected expression of keratin 19 and nuclear binding sites for galectin-1 in adult cells migrating from the hair follicle, where cells expressing keratin 19 are located in the bulge region. The results were compared with the expression of both markers in cells adhering from suspension prepared from the interfollicular epidermis without keratin-19-positive cells and with nuclear binding sites for galectin-1. The results were compared with data from basal cell carcinomas. All cells were analyzed concerning size, as it is known that cell diameter influences the clonogenic potential of keratinocytes. The major result of this study is the observation of transient expression of keratin 19 and nuclear galectin-1 binding sites in originally negative interfollicular epidermal cells induced by adhesion. These cells were very small in size, similar to basal cells of the interfollicular epidermis or the bulge region of the hair follicle. The influence of the suspension regimen on beta1-integrin expression, cell diameter and growth was also monitored. A population of cells highly positive for beta1 integrin of the same diameter as keratin-19-positive cells insensitive to induction of terminal differentiation by lack of anchorage was characterized. Cells of the same size were also observed in the keratin-19-positive cells of basal cell carcinomas. In conclusion, the expression of poor levels of differentiation induced by cell adhesion is transient. Also, keratin 19 expression should not be exclusively regarded as a marker of stem cell activity.

• MeSH
• antigeny CD29 analýza   MeSH
• bazocelulární karcinom metabolismus   patologie   MeSH
• buněčná adheze MeSH
• buněčné kultury MeSH
• časové faktory MeSH
• epidermální buňky MeSH
• epidermis chemie   MeSH
• financování organizované MeSH
• galektin 1 analýza   MeSH
• galektin 3 analýza   MeSH
• keratinocyty cytologie   chemie   MeSH
• keratiny analýza   MeSH
• kultivované buňky MeSH
• lidé MeSH
• nádory kůže metabolismus   patologie   MeSH
• pohyb buněk MeSH
• proteiny intermediálních filament analýza   MeSH
• vlasový folikul cytologie   chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• srovnávací studie MeSH

The transient receptor potential channel A1 (TRPA1) is unique among ion channels of higher vertebrates in that it harbors a large ankyrin repeat domain. The TRPA1 channel is expressed in the inner ear and in nociceptive neurons. It is involved in hearing as well as in the perception of pungent and irritant chemicals. The ankyrin repeat domain has special mechanical properties, which allows it to function as a soft spring that can be extended over a large range while maintaining structural integrity. A calcium-binding site has been experimentally identified within the ankyrin repeats. We built a model of the N-terminal 17 ankyrin repeat structure, including the calcium-binding EF-hand. In our simulations we find the calcium-bound state to be rigid as compared to the calcium-free state. While the end-to-end distance can change by almost 50% in the apo form, these fluctuations are strongly reduced by calcium binding. This increase in stiffness that constraints the end-to-end distance in the holo form is predicted to affect the force acting on the gate of the TRPA1 channel, thereby changing its open probability. Simulations of the transmembrane domain of TRPA1 show that residue N855, which has been associated with familial episodic pain syndrome, forms a strong link between the S4-S5 connecting helix and S1, thereby creating a direct force link between the N-terminus and the gate. The N855S mutation weakens this interaction, thereby reducing the communication between the N-terminus and the transmembrane part of TRPA1.

• MeSH
• ankyrinová repetice fyziologie   MeSH
• kationtové kanály TRP chemie   fyziologie   MeSH
• lidé MeSH
• molekulární modely MeSH
• motivy EF-ruky fyziologie   MeSH
• proteiny nervové tkáně chemie   fyziologie   MeSH
• simulace molekulární dynamiky MeSH
• vápník metabolismus   MeSH
• vápníkové kanály chemie   fyziologie   MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

In mammals, double-stranded RNA (dsRNA) can mediate sequence-specific RNA interference, activate sequence-independent interferon response, or undergo RNA editing by adenosine deaminases. We showed that long hairpin dsRNA expression had negligible effects on mammalian somatic cells--expressed dsRNA was slightly edited, poorly processed into siRNAs, and it did not activate the interferon response. At the same time, we noticed reduced reporter expression in transient co-transfections, which was presumably induced by expressed dsRNA. Since transient co-transfections are frequently used for studying gene function, we systematically explored the role of expressed dsRNA in this silencing phenomenon. We demonstrate that dsRNA expressed from transiently transfected plasmids strongly inhibits the expression of co-transfected reporter plasmids but not the expression of endogenous genes or reporters stably integrated in the genome. The inhibition is concentration-dependent, it is found in different cell types, and it is independent of transfection method and dsRNA sequence. The inhibition occurs at the level of translation and involves protein kinase R, which binds the expressed dsRNA. Thus, dsRNA expression represents a hidden danger in transient transfection experiments and must be taken into account during interpretation of experimental results.

• MeSH
• buňky 3T3 MeSH
• dvouvláknová RNA metabolismus   MeSH
• HEK293 buňky MeSH
• HeLa buňky MeSH
• imunoprecipitace MeSH
• lidé MeSH
• malá interferující RNA genetika   MeSH
• myši MeSH
• plazmidy genetika   MeSH
• protein-serin-threoninkinasy metabolismus   MeSH
• průtoková cytometrie MeSH
• regulace genové exprese genetika   MeSH
• reportérové geny genetika   MeSH
• transfekce metody   MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH