Mapping of alpha-neo-endorphin- and neurokinin B-immunoreactivity in the human brainstem

We have studied the distribution of alpha-neo-endorphin- or neurokinin B-immunoreactive fibres and cell bodies in the adult human brainstem with no prior history of neurological or psychiatric disease. A low density of alpha-neo-endorphin-immunoreactive cell bodies was only observed in the medullary central gray matter and in the spinal trigeminal nucleus (gelatinosa part). Alpha-neo-endorphin-immunoreactive fibres were moderately distributed throughout the human brainstem. A high density of alpha-neo-endorphin-immunoreactive fibres was found only in the solitary nucleus (caudal part), in the spinal trigeminal nucleus (caudal part), and in the gelatinosa part of the latter nucleus. Neurokinin B-immunoreactive cell bodies (low density) were found in the periventricular central gray matter, the reticular formation of the pons and in the superior colliculus. The distribution of the neurokinin-immunoreactive fibres was restricted. In general, for both neuropeptides the density of the immunoreactive fibres was low. In the human brainstem, the proenkephalin system was more widely distributed than the prodynorphin system, and the preprotachykinin A system (neurokinin A) was more widely distributed than the preprotachykinin B system (neurokinin B).     

Synthetic non-classical luminescence generation by enhanced silica nanophotonics based on nano-bio-FRET

Fluorescent silica nanoparticles (NPs–(SiO₂–Fluo)) were synthesized based on the classical Störber method for cyanobacteria labelling. Modified mono-coloured SiO₂ NPs with fluorescein (Fl) and rhodamine B (RhB) were obtained (NPs–(SiO₂–Fl) and NPs–(SiO₂–RhB)). Moreover, multi-coloured SiO₂ NPs, via the incorporation of both emitters (NPs–(SiO₂–RhB–Fl)), were tuned for optimal emissions and the biodetection of cyanobacteria. NPs–(SiO₂–Fl) and NPs–(SiO₂–RhB–Fl) were optimized for detection via laser fluorescence microscopy and in-flow cytometry with laser excitation and fluorescence detection. By TEM, homogeneous SiO₂ NPs of 180.0 nm in diameter were recorded. These sizes were slightly increased due to the covalent linking incorporation of fluorescent dye emitters to 210.0 nm with mono-coloured fluorescent modified amine-organosilanes, and to 340.0 nm in diameter with multi-coloured dye incorporation. NPs–(SiO₂–Fluo) showed variable emission depending on the dye emitter concentration, quantum yield and applied luminescent pathway. Thus, mono-coloured NPs–(SiO₂–Fl) and NPs–(SiO₂–RhB) showed diminished emissions in comparison to multi-coloured NPs–(SiO₂–RhB–Fl). This enhancement was explained by fluorescence resonance energy transfer (FRET) between Fl as a fluorescent energy donor and RhB as an energy acceptor produced within the nanoarchitecture, produced only in the presence of both fluorophores with the appropriate laser excitation of the energy donor. The depositions of the nano-emitters on cyanobacteria by non-covalent interactions were observed by TEM and laser fluorescence microscopy. For multi-coloured NPs–(SiO₂–RhB–Fl) labelling, bio-FRET was observed between the emission of the nano-labellers and the natural fluorophores from the cyanobacteria that quenched the emission of the whole nano-biostructure in comparison to mono-coloured NPs–(SiO₂–Fl) labelling. This fact was explained and discussed in terms of different fluorescence energy transfer from the nanolabellers towards different natural chromophore coupling. In the presence of NPs–(SiO₂–RhB–Fl) and NPs–(SiO₂–RhB), the emission was coupled with lower quantum yield chromophores; while upon the application of NPs–(SiO₂–Fl), it was coupled with higher quantum yield chromophores. In this manner, for enhanced luminescent nanoplatform tracking, the multi-coloured NPs–(SiO₂–RhB–Fl) showed improved properties; but more highly luminescent bio-surfaces were generated with mono-coloured NPs–(SiO₂–Fl) that permitted faster cyanobacteria detection and counting by laser fluorescence microscopy, and by in-flow cytometry with laser excitation and fluorescence detection.

Fluorescent silica nanophotonics for cyanobacteria labelling.     

Validity of model approximations for receptor-ligand kinetics in nuclear medicine

An appropriate mathematical model is required for quantitative analysis of high affinity radioligands as direct or surrogate probes to measure receptor distribution, affinity, concentration, binding potential, and endogenous or exogenous ligand occupancy levels. For studies with positron emission tomography (PET) or single photon emission computed tomography (SPECT), the receptor-ligand compartment model has been well established and widely used. This pharmacokinetic model is represented mathematically by a set of nonlinear ordinary differential equations. Variations of models for PET and SPECT account for radioactive decay differently. These are not equivalent and entail assumptions or approximations that may be not appreciated. In this study, a general form of the model is presented and compared with others with various approximations, which are valid only under specific conditions. The various approximate formulations were analytically compared to the exact model to identify the terms that were neglected in the approximate formulations. The extent to which the approximations impact the model solutions was assessed by computer simulations based on numerical solutions to each set of equations. Specifically, each model formulation was tested using three simulated injection protocols representing a typical PET experiment, a typical SPECT experiment, and an extreme experiment where both the injected activity and the specific activity were very high. No significant differences were found among the output of the three model formulations when the PET and SPECT injection protocols were tested. The only conditions that produced significant differences occurred when the specific activity and the administered activity were simultaneously very high. These conditions, however, have little practical relevance to experimentally achievable conditions due to radiation dose and specific activity of radiopharmaceuticals     

A DNA aptamer population specifically detects Leishmania infantum H2A antigen

Aptamers are short single-stranded DNA or RNA oligonucleotides that are selected in vitro by their affinity and specificity for the target. Binding is a consequence of the particular tertiary structure that they are able to acquire, depending on their sequence. Parasites of the genus Leishmania belongs to the lower eukaryote order Kinetoplastida that causes leishmaniosis in man and animals. Histone genes in Leishmania are of considerable interest because these flagellates do not condense their chromatin during mitosis. Thus, the study of the structural features of histones has been considered of particular interest and, as a result, in recent years a great number of histone genes have been characterized in trypanosomatids. Histones are extremely conserved proteins, reflecting their apparent universality of function. Sequence similarity of kinetoplastid core histones those of higher eukaryotes is found predominantly in the globular region with high sequence divergences in the N- and in the C-terminal domains. These divergences indicate that they may be potential diagnostic and/or therapeutics targets. We have successfully isolated a pool of DNA sequences, named SELH2A, which specifically binds to Leishmania infantum H2A. When tested in an enzyme-linked oligonucleotide assay, slot blot and Western blot analysis, the aptamer pool exhibited specificity in its ability to bind only to H2A antigen but not to other proteins from L. infantum including other histones. Thus, it appears that this novel anti-H2A aptamer population may be of potential application as a diagnostic system for leishmaniosis.