Concept of energy in biological systems and the effects of irradiations of low energies on enzyme-substrate systems

The recent mathematical formalization of the concepts of matter and extrinsical energy, which are used for the relational representation of biological systems, is employed in the analysis of the important experimental discoveries of Comorosanet al. related to low energy electromagnetic irradiations on enzyme substrates. By means of the present analysis one of the properties inherent to the experimental phenomena is more precisely exposed, and theoretical developments corresponding to “energetical evolutions” in a biological system (Leguizamón, 1976) may now have an experimental basis. Important limitations are introduced for the validity of the commutativity and associativity of cartesian product of sets, when they represent matter and its linked extrinsical energy. In connection with this last aspect, new important knowledge is obtained for the relational mathematical representation of biological systems.     

Evolution of the nucleotide sequence of influenza virus RNA segment 7 during drift of the H3N2 subtype

The complete genetic information contained in the influenza virus RNA segment 7 of the A/Bangkok/ $\text{1}\text{79}$ (H3N2) strain has been cloned by in vitro synthesis of the complementary dsDNA and its insertion into plasmid pBR322. The nucleotide sequence of the viral RNA segment has been determined from the cDNA insert. It is 1027 nucleotides long, and contains two open reading frames, as shown for other influenza virus strains. When compared with the previously published sequence for the A/Udorn/72 (H3N2) strain, 15 nucleotide exchanges are observed, most of them silent mutations, and only two causing amino acid changes in each of the M1 and M2 protein sequences.     

Tyrosine transport by membrane vesicles isolated from rat brain

Tyrosine uptake by membrane vesicles derived from rat brain has been investigated. The uptake is dependent on an Na⁺ gradient ([$\text{Na}{}^{\mathrm{+}}\text{]}\text{outside}\text{> [}\text{Na}{}^{\mathrm{+}}\text{]}\text{inside}$). The uptake is transport into an osmotically active space and not a binding artifact as indicated by the effect of increasing the medium osmolarity. The process is stimulated by a membrane potential (negative inside) as demonstrated by the effect of the ionophores valinomycin and carbonyl cyanide $\text{m-}\text{chlorophenylhydrazone}$ and anions with different permeabilities. Kinetic data show that tyrosine is accumulated by two systems with different affinities. Tyrosine uptake is inhibited by the presence of phenylalanine and tryptophan.     

Differential Modulation of TREM2 Protein during Postnatal Brain Development in Mice

During postnatal development, microglia, the resident innate immune cells of the central nervous system are constantly monitoring the brain parenchyma, cleaning the cell debris, the synaptic contacts overproduced and also maintaining the brain homeostasis. In this context, the postnatal microglia need some control over the innate immune response. One such molecule recently described to be involved in modulation of immune response is TREM2 (triggering receptor expressed on myeloid cells 2). Although some studies have observed TREM2 mRNA in postnatal brain, the regional pattern of the TREM2 protein has not been described. We therefore characterized the distribution of TREM2 protein in mice brain from Postnatal day (P) 1 to 14 by immunostaining. In our study, TREM2 protein was expressed only in microglia/macrophages and is developmentally downregulated in a region-dependent manner. Its expression persisted in white matter, mainly in caudal corpus callosum, and the neurogenic subventricular zone for a longer time than in grey matter. Additionally, the phenotypes of the TREM2+ microglia also differ; expressing CD16/32, MHCII and CD86 (antigen presentation markers) and CD68 (phagocytic marker) in different regions as well as with different intensity till P7. The mannose receptor (CD206) colocalized with TREM2 only at P1–P3 in the subventricular zone and cingulum, while others persisted at low intensities till P7. Furthermore, the spatiotemporal expression pattern and characterization of TREM2 indicate towards its other plausible roles in phagocytosis, progenitor’s fate determination or microglia phenotype modulation during postnatal development. Hence, the increase of TREM2 observed in pathologies may recapitulate their function during postnatal development, as a better understanding of this period may open new pathway for future therapies.     

The novel MER transposon-derived miRNAs in human genome

MicroRNAs (miRNAs) are small RNA molecules (~ 20–30 nucleotides) that generally act in gene silencing and translational repression through the RNA interference pathway. They generally originate from intergenic genomic regions, but some are found in genomic regions that have been characterized such as introns, exons, and transposable elements (TE). To identify the miRNAs that are derived from palindromic MERs, we analyzed MER paralogs in human genome. The structures of the palindromic MERs were similar to the hairpin structure of miRNA in humans. Three miRNAs derived from MER96 located on chromosome 3, and MER91C paralogs located on chromosome 8 and chromosome 17 were identified in HeLa, HCT116, and HEK293 cell lines. The interactions between these MER-derived miRNAs and AGO1, AGO2, and AGO3 proteins were validated by immunoprecipitation assays. The data suggest that miRNAs derived from transposable elements could widely affect various target genes in the human genome.     

Characterization of a Novel Porcine Parvovirus Tentatively Designated PPV5

A new porcine parvovirus (PPV), provisionally designated as PPV5, was identified in U.S. pigs. Cloning and sequencing from a circular or head-to-tail concatemeric array revealed that the PPV5 possesses the typical genomic organization of parvoviruses with two major predicted open reading frames (ORF1 and ORF2), and is most closely related to PPV4 with overall genomic identities of 64.1–67.3%. The amino acid identities between PPV5 and PPV4 were 84.6%–85.1% for ORF1 and 54.0%–54.3% for ORF2. Unlike PPV4, but similar to bovine parvovirus 2 (BPV2), PPV5 lacks the additional ORF3 and has a much longer ORF2. Moreover, the amino acid sequences of ORF1 and ORF2 of BPV2 showed higher homologies to PPV5 than to PPV4. The conserved motifs of the Ca²⁺ binding loop (YXGXG) and the catalytic center (HDXXY) of phospholipase A₂ (PLA₂) were identified in VP1 (ORF2) of PPV5, as well as in BPV2, but were not present in PPV4. Phylogenetic analyses revealed that PPV5, PPV4 and BPV2 form a separate clade different from the genera Parvovirus and Bocavirus. Further epidemiologic investigations of PPV4 and PPV5 in U.S. pigs of different ages indicated a slightly higher prevalence for PPV5 (6.6%; 32/483) compared to PPV4 (4.1%; 20/483), with detection of concurrent PPV4 and PPV5 in 15.6% (7/45) of lungs of infected pigs. Evidence for potential vertical transmission or association with reproductive failure was minimal for both PPV4 and PPV5. The high similarity to PPV4 and the lack of ORF3 may suggest PPV5 is an intermediate of PPV4 during the evolution of parvoviruses in pigs.     

Preparation and characterization of bosentan monohydrate/ε-polycaprolactone nanoparticles obtained by electrospraying

The electrospraying technique provides nano and microparticles that can be used as drug delivery systems. The aims of this study were, firstly, to optimize the influent parameters of electrospraying for the manufacture of a Bosentan (BOS) nanoparticulate platform, and secondly, to evaluate its physicochemical properties and in vitro biopharmaceutical behavior. Particles were characterized by scanning electron microscopy (SEM), powder X-ray diffraction (PXRD), differential scanning calorimetry (DSC), thermogravimetry (TG) and Fourier transformed Infrared spectroscopy (FTIR). Drug loading, encapsulation efficiency and kinetic dissolution were determined. Additionally, Bosentan release assays at 24 and 72 h were performed in vitro to evaluate biopharmaceutical properties of nano-scaffolds by diffusion technique through dialysis bag. The nanostructures had heterogeneous sizes predominantly smaller than 550 nm and they were semicrystalline according to PXRD, indicating a partial amorphization of BOS during the encapsulation in the polymer matrix. FT-IR and DSC showed an absence of chemical interactions between BOS and ε-Polycaprolactone (PCL), suggesting that both components behaved as a physical mixture in these particles. The drug loading was 25.98%, and the encapsulation efficiency was 58.51%. Additionally, the release assays showed an extended and controlled release of BOS, in comparison to non-encapsulated BOS. These data also showed to fit with the Cubic Root kinetic dissolution. As a conclusion, we demonstrate that the use of electrospraying for the manufacture of BOS (or similar drugs) controlled release nanoplatforms would represent an interesting contribution in the development of new therapeutic alternatives for the treatment of pathologies such as pulmonary hypertension and other related diseases. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2748, 2019.     

Chromatin reorganization during spermiogenesis of the mollusc Thais hemostoma (Muricidae): implications for sperm nuclear morphogenesis in cenogastropods

Thais is a cenogastropod mollusc belonging to the Muricidae family. The sperm nuclear morphogenesis of Thais develops in two well-defined and peculiar steps. In the first one, the round early spermatidyl nucleus is penetrated by an endonuclear channel, which arranges as a helix at the inner nuclear surface and organizes the condensing chromatin all around. In the second step, the spiral channel stretches, dragging along the associated chromatin and leading to a definitive cylinder-shaped sperm nucleus. Simultaneously with these changes in nuclear shape, the chromatin is sequentially organized in granules, fibres, lamellae, and, finally, in a very condensed structure, whereas the spermiogenic DNA-associated proteins become more basic and simple. The sperm nucleus contains a small group of protamines consisting of only four types of amino acid (lysine, arginine, glycine, and serine). The most remarkable fact on nuclear spermiogenesis in Thais is that, whereas the chromatin condensation process, the nuclear proteins, and the final shape of sperm nucleus are very similar to those in other muricidae studied, the pathway of nuclear morphogenesis is completely different. We propose an independent genetic control for those two spermiogenic events (chromatin condensation and nucleomorphogenesis). Finally we discuss briefly the main traits of nucleomorphogenesis of muricid molluscs.     

Notes on the combined use of V-VIP and DAB peroxidase substrates for the detection of colocalising antigens

 The purpose of the present report was to investigate to what extent the new peroxidase substrate Vector VIP (V-VIP) can be used in combination with DAB chromogen for the unequivocal and permanent detection of colocalising antigens within a single neurone, according to a two-colour paradigm. With this aim, re-trograde tract-tracing with cholera toxin B subunit (CTB) or fluoro-gold (FG) was performed to disclose individual, identified subpopulations of neurones in the primate substantia nigra projecting to the caudate nucleus or to the putamen, respectively. Each tracer was detected by means of a PAP procedure and finally stained brown using DAB as a chromogen. Subsequently, both series of sections were processed for the immunocytochemical detection of tyrosine hydroxylase (TH). TH-immunoreactive neurones were stained purple with the peroxidase substrate V-VIP. As a result of the present procedure, several cell bodies of projection neurones, stained brown, can easily be identified within the primate substantia nigra. Some of these neurones additionally displayed purple TH immunoreaction product located in the neuronal dendrites. By contrast, CTB- or FG-unlabelled neurones only show the typical purple precipitate that belongs to V-VIP substrate, both in the cell body as well as in the dendrites. Accepted: 20 January 1999