The endothelin axis (endothelins and their receptors) is strongly involved in physiological and pathological processes. ET-1 plays a crucial role in particular in tumor diseases. Endothelin-1 receptors (ET(A) and ET(B)) are deregulated and overexpressed in several tumors such as melanoma and glioma. We studied the binding of 24 monoclonal antibodies directed against human ET(B) receptors (hET(B)) to different melanoma cell lines. Few of these mAbs bound to all the melanoma cell lines. One of them, rendomab B49, bound to ET(B) receptors expressed at the surface of human glioma stem cells. More recently, we produced new antibodies directed against human ET(A) receptor (hET(A)). Several antibodies have been isolated and have been screened on different tumoral cells lines. As for the mAbs directed against the hET(B) receptor only some of new antibodies directed against ET(A) receptor are capable to bind the human tumoral cell lines. Rendomab A63 directed against hET(A) is one of them. We report the specificity and binding properties of these mAbs and consider their potential use in diagnosis by an in vivo imaging approach.
- MeSH
- antagonisté endotelinového receptoru A aplikace a dávkování metabolismus MeSH
- CHO buňky MeSH
- Cricetulus MeSH
- endotelin-1 genetika metabolismus MeSH
- křečci praví MeSH
- lidé MeSH
- monoklonální protilátky aplikace a dávkování metabolismus MeSH
- myši nahé MeSH
- myši MeSH
- nádorové buněčné linie MeSH
- receptor endotelinu A genetika metabolismus MeSH
- receptor endotelinu B genetika metabolismus MeSH
- sekvence aminokyselin MeSH
- vazba proteinů fyziologie MeSH
- xenogenní modely - testy antitumorózní aktivity metody MeSH
- zvířata MeSH
- Check Tag
- křečci praví MeSH
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Neurones in the supraoptic nucleus (SON) of the hypothalamus possess intrinsic osmosensing mechanisms, which are lost in transient receptor potential vanilloid 1 (Trpv1)-knock-out mice. The molecular nature of the osmosensory mechanism in SON neurones is believed to be associated with the N-terminal splice variant of Trpv1, although their entire molecular structures have not been hitherto identified. In this study, we sought for TRPV1-related molecules and their function in the rat SON. We performed RT-PCR and immunohistochemistry to detect TRPV1-related molecules in the SON, and patch-clamp and imaging of the cytosolic Ca(2+) concentration ([Ca(2+)]i) to measure responses to osmolality changes and TRPV-related drugs in acutely dissociated SON neurones of rats. RT-PCR analysis revealed full-length Trpv1 and a new N-terminal splice variant, Trpv1_SON (LC008303) in the SON. Positive immunostaining was observed using an antibody against the N-terminal portion of TRPV1 in arginine vasopressin (AVP)-immunoreactive neurones, but not in oxytocin (OT)-immunoreactive neurones. Approximately 20% of SON neurones responded to mannitol (50 mM) with increased action potential firing, inward currents, and [Ca(2+)]i mobilization. Mannitol-induced responses were observed in AVP neurones isolated from AVP-eGFP transgenic rats and identified by GFP fluorescence, but not in OT neurones isolated from OT-mRFP transgenic rats and identified by RFP fluorescence. The mannitol-induced [Ca(2+)]i responses were reversibly blocked by the non-selective TRPV antagonist, ruthenium red (10 μM) and the TRPV1 antagonists, capsazepine (10 μM) and BCTC (10 μM). Although the TRPV1 agonist, capsaicin (100 nM) evoked no response at room temperature, it triggered cationic currents and [Ca(2+)]i elevation when the temperature was increased to 36°C. These results suggest that AVP neurones in the rat SON possess functional full-length TRPV1. Moreover, differences between the responses to capsaicin or hyperosmolality obtained in rat SON neurones and those obtained from dorsal root ganglion neurones or TRPV1-expressing cells indicate that the osmoreceptor expressed in the SON may be a heteromultimer in which TRPV1 is co-assembled with some other, yet unidentified, molecules.
- MeSH
- akční potenciály účinky léků MeSH
- HEK293 buňky MeSH
- kapsaicin analogy a deriváty farmakologie MeSH
- kationtové kanály TRPV agonisté genetika metabolismus MeSH
- krysa rodu rattus MeSH
- kultivované buňky MeSH
- lidé MeSH
- mannitol farmakologie MeSH
- neurony cytologie metabolismus MeSH
- nucleus supraopticus metabolismus MeSH
- osmolární koncentrace MeSH
- oxytocin farmakologie MeSH
- potkani transgenní MeSH
- potkani Wistar MeSH
- pyraziny farmakologie MeSH
- pyridiny farmakologie MeSH
- teplota MeSH
- vápníková signalizace účinky léků MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- lidé MeSH
- mužské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
There is still a considerable need for development of new tools and methods detecting specific viral proteins for the diagnosis and pathogenesis study of the Yellow fever virus (YFV). This study aimed to develop and characterize polyclonal peptide antisera for detection of YFV-C and -NS1 proteins. The antisera were used further to investigate NS1 protein expression during YFV infection in mammalian cells. YFV target proteins were detected by all antisera in western blot and immunofluorescence assays. No cross-reactivity was observed with Dengue virus, West Nile virus, Tick-borne encephalitis virus and Japanese encephalitis virus. Nuclear localization of the YFV-C protein was demonstrated for the first time. Experiments investigating NS1 expression suggested a potential use of the YFV-NS1 antisera for development of diagnostic approaches targeting the secreted form of the NS1 protein. The antisera described in this study offer new possibilities for use in YFV research and for the development of novel diagnostic tests.
- MeSH
- antigeny virové analýza imunologie metabolismus MeSH
- buněčné jádro chemie virologie MeSH
- Cercopithecus aethiops MeSH
- diagnostické testy rutinní metody MeSH
- fluorescenční protilátková technika MeSH
- králíci MeSH
- morčata MeSH
- peptidy izolace a purifikace metabolismus MeSH
- protilátky virové imunologie izolace a purifikace metabolismus MeSH
- senzitivita a specificita MeSH
- Vero buňky MeSH
- virologie metody MeSH
- virové nestrukturální proteiny analýza imunologie metabolismus MeSH
- virové plášťové proteiny analýza imunologie metabolismus MeSH
- virus žluté zimnice imunologie izolace a purifikace MeSH
- western blotting MeSH
- zkřížené reakce MeSH
- zvířata MeSH
- Check Tag
- králíci MeSH
- morčata MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
