Simulating the embolization of blood vessels using magnetic microparticles and acupuncture needle in a magnetic field

Computer models were developed to simulate the capture and subsequent deposition of magnetic microparticles (MMPs) in a blood vessel adjacent to a ferromagnetic wire (e.g., acupuncture needle) magnetized by a uniform external magnetic field. Process parameter conditions were obtained to enable optimal capture of MMPs into the deposit. It was found that the maximum capture distance of the MMPs was within 0.5-2.0 mm when the particles were superparamagnetic and had large size (>1.0 microm) and relative large flow rates (2.5-5.0 cm/s) as in a healthy artery. It was also found that the deposits were asymmetrical and that their size was between 1.0 and 2.0 mm. For the case of lower flow rates as can be found in a tumor (<1.0 mm/s) and using small magnetite particles (0.25-2.0 microm) the maximum capture distance was larger, ranging between approximately 0.5 and 6.4 mm, depending on the blood flow rate, the radius of wire, and particle clustering. The range of embolization (deposition) in this later case was between 0.5 and 5.9 mm. The potential of this technique to generate MMPs deposits to embolize blood vessels inhibiting the blood supply and thus facilitating necrosis of tumors located deep within the patient (3-7 cm) is discussed.     

The carboxy-terminal fragment of nucleolin interacts with the nucleocapsid domain of retroviral gag proteins and inhibits virion assembly

A yeast two-hybrid screen for cellular proteins that interact with the murine leukemia virus (MuLV) Gag protein resulted in the identification of nucleolin, a host protein known to function in ribosome assembly. The interacting fusions contained the carboxy-terminal 212 amino acids of nucleolin [Nuc(212)]. The nucleocapsid (NC) portion of Gag was necessary and sufficient to mediate the binding to Nuc(212). The interaction of Gag with Nuc(212) could be demonstrated in vitro and was manifested in vivo by the NC-dependent incorporation of Nuc(212) inside MuLV virions. Overexpression of Nuc(212), but not full-length nucleolin, potently and specifically blocked MuLV virion assembly and/or release. A mutant of MuLV, selected to specifically disrupt the binding to Nuc(212), was found to be severely defective for virion assembly. This mutant harbors a single point mutation in capsid (CA) adjacent to the CA-NC junction, suggesting a role for this region in Moloney MuLV assembly. These experiments demonstrate that selection for proteins that bind assembly domain(s) can yield potent inhibitors of virion assembly. These experiments also raise the possibility that a nucleolin-Gag interaction may be involved in virion assembly.     

No evidence of inhibition of horizontal gene transfer by CRISPR–Cas on evolutionary timescales

The CRISPR (clustered, regularly, interspaced, short, palindromic repeats)–Cas (CRISPR-associated genes) systems of archaea and bacteria provide adaptive immunity against viruses and other selfish elements and are believed to curtail horizontal gene transfer (HGT). Limiting acquisition of new genetic material could be one of the sources of the fitness cost of CRISPR–Cas maintenance and one of the causes of the patchy distribution of CRISPR–Cas among bacteria, and across environments. We sought to test the hypothesis that the activity of CRISPR–Cas in microbes is negatively correlated with the extent of recent HGT. Using three independent measures of HGT, we found no significant dependence between the length of CRISPR arrays, which reflects the activity of the immune system, and the estimated number of recent HGT events. In contrast, we observed a significant negative dependence between the estimated extent of HGT and growth temperature of microbes, which could be explained by the lower genetic diversity in hotter environments. We hypothesize that the relevant events in the evolution of resistance to mobile elements and proclivity for HGT, to which CRISPR–Cas systems seem to substantially contribute, occur on the population scale rather than on the timescale of species evolution.     

Amplification of a Cytochrome P450 Gene Is Associated with Resistance to Neonicotinoid Insecticides in the Aphid Myzus persicae

The aphid Myzus persicae is a globally significant crop pest that has evolved high levels of resistance to almost all classes of insecticide. To date, the neonicotinoids, an economically important class of insecticides that target nicotinic acetylcholine receptors (nAChRs), have remained an effective control measure; however, recent reports of resistance in M. persicae represent a threat to the long-term efficacy of this chemical class. In this study, the mechanisms underlying resistance to the neonicotinoid insecticides were investigated using biological, biochemical, and genomic approaches. Bioassays on a resistant M. persicae clone (5191A) suggested that P450-mediated detoxification plays a primary role in resistance, although additional mechanism(s) may also contribute. Microarray analysis, using an array populated with probes corresponding to all known detoxification genes in M. persicae, revealed constitutive over-expression (22-fold) of a single P450 gene (CYP6CY3); and quantitative PCR showed that the over-expression is due, at least in part, to gene amplification. This is the first report of a P450 gene amplification event associated with insecticide resistance in an agriculturally important insect pest. The microarray analysis also showed over-expression of several gene sequences that encode cuticular proteins (2–16-fold), and artificial feeding assays and in vivo penetration assays using radiolabeled insecticide provided direct evidence of a role for reduced cuticular penetration in neonicotinoid resistance. Conversely, receptor radioligand binding studies and nucleotide sequencing of nAChR subunit genes suggest that target-site changes are unlikely to contribute to resistance to neonicotinoid insecticides in M. persicae.     

Altered expression of claudin family proteins in Alzheimer’s disease and vascular dementia brains

Claudins (Cls) are a multigene family of transmembrane proteins with different tissue distribution, which have an essential role in the formation and sealing capacity of tight junctions (TJs). At the level of the blood–brain barrier (BBB), TJs are the main molecular structures which separate the neuronal milieu from the circulatory space, by a restriction of the paracellular flow of water, ions and larger molecules into the brain. Different studies suggested recently significant BBB alterations in both vascular and degenerative dementia types. In a previous study we found in Alzheimer’s disease (AD) and vascular dementia (VaD) brains an altered expression of occludin, a molecular partner of Cls in the TJs structure. Therefore in this study, using an immunohistochemical approach, we investigated the expression of Cl family proteins (Cl‐2, Cl‐5 and Cl‐11) in frontal cortex of aged control, AD and VaD brains. To estimate the number of Cl‐expressing cells, we applied a random systematic sampling and the unbiased optical fractionator method. We found selected neurons, astrocytes, oligodendrocytes and endothelial cells expressing Cl‐2, Cl‐5 and Cl‐11 at detectable levels in all cases studied. We report a significant increase in ratio of neurons expressing Cl‐2, Cl‐5 and Cl‐11 in both AD and VaD as compared to aged controls. The ratio of astrocytes expressing Cl‐2 and Cl‐11 was significantly higher in AD and VaD as compared to aged controls. The ratio of oligodendrocytes expressing Cl‐11 was significantly higher in AD and the ratio of oligodendrocytes expressing Cl‐2 was significantly higher in VaD as compared to aged controls. Within the cerebral cortex, Cls were selectively expressed by pyramidal neurons, which are the ones responsible for cognitive processes and affected by AD pathology. Our findings suggest a new function of Cl family proteins which might be linked to response to cellular stress.     

Interaction of inorganic mercury with CoA-SH and acyl-CoAs

Sulfur containing biomolecules are involved in complexes with mercury. CoA is an important cofactor for many enzymes involved in metabolic processes. Fatty acyl-CoA-thioesters, substrates of mitochondrial ß-oxidation, are sulfur containing compounds and potential mercury ligands. The CoA-Hg²⁺ complex can be easily assessed by UV-Vis spectroscopy or indirectly by antibacterial tests that reconfirmed the protective role of CoA on E. coli. The characteristics of these complexes were determined by means of FTIR spectroscopy. The reverse phase liquid chromatography combined with electrospray ionization tandem mass spectrometry was used for detection of the side-product that resulted through the cleavage of thioesters in the presence of mercury. An unexpected result was the detection of octathioic acid as a product. Our study shows that mitochondrial β-oxidation can be affected by thioesters depletion assisted by Hg²⁺. The GC-MS technique could be used to detect some possible mitochondrial injuries due to the heavy metal ions.     

Trends and barriers to lateral gene transfer in prokaryotes

Gene acquisition by lateral gene transfer (LGT) is an important mechanism for natural variation among prokaryotes. Laboratory experiments show that protein-coding genes can be laterally transferred extremely fast among microbial cells, inherited to most of their descendants, and adapt to a new regulatory regime within a short time. Recent advance in the phylogenetic analysis of microbial genomes using networks approach reveals a substantial impact of LGT during microbial genome evolution. Phylogenomic networks of LGT among prokaryotes reconstructed from completely sequenced genomes uncover barriers to LGT in multiple levels. Here we discuss the kinds of barriers to gene acquisition in nature including physical barriers for gene transfer between cells, genomic barriers for the integration of acquired DNA, and functional barriers for the acquisition of new genes.