Fluorescently labeled differentiating myelopeptide-4: Specific binding to and penetration into target cells

Myelopeptide-4 (MP-4) (Phe-Arg-Pro-Arg-Ile-Met-Thr-Pro), inducing the terminal differentiation of HL-60 leukemia cells, was labeled with fluorescein isothiocyanate. The specific binding of this modified peptide to the surface of HL-60 cells and its ability to penetrate into the cells were studied. It was shown by cytometry and confocal microscopy to be bound on the HL-60 cell surface, to penetrate into their cytoplasm, and finally to concentrate around the cell nucleus. These phenomena are probably necessary for the exhibition of MP-4 differentiating activity.     

Effect of self-association on the structural organization of partially folded proteins: inactivated actin

The propensity to associate or aggregate is one of the characteristic properties of many nonnative proteins. The aggregation of proteins is responsible for a number of human diseases and is a significant problem in biotechnology. Despite this, little is currently known about the effect of self-association on the structural properties and conformational stability of partially folded protein molecules. G-actin is shown to form equilibrium unfolding intermediate in the vicinity of 1.5 M guanidinium chloride (GdmCl). Refolding from the GdmCl unfolded state is terminated at the stage of formation of the same intermediate state. An analogous form, known as inactivated actin, can be obtained by heat treatment, or at moderate urea concentration, or by the release of Ca(2+). In all cases actin forms specific associates comprising partially folded protein molecules. The structural properties and conformational stability of inactivated actin were studied over a wide range of protein concentrations, and it was established that the process of self-association is rather specific. We have also shown that inactivated actin, being denatured, is characterized by a relatively rigid microenvironment of aromatic residues and exhibits a considerable limitation in the internal mobility of tryptophans. This means that specific self-association can play an important structure-forming role for the partially folded protein molecules.     

Male and female brain evolution is subject to contrasting selection pressures in primates

The claim that differences in brain size across primate species has mainly been driven by the demands of sociality (the "social brain" hypothesis) is now widely accepted. Some of the evidence to support this comes from the fact that species that live in large social groups have larger brains, and in particular larger neocortices. Lindenfors and colleagues (BMC Biology 5:20) add significantly to our appreciation of this process by showing that there are striking differences between the two sexes in the social mechanisms and brain units involved. Female sociality (which is more affiliative) is related most closely to neocortex volume, but male sociality (which is more competitive and combative) is more closely related to subcortical units (notably those associated with emotional responses). Thus different brain units have responded to different selection pressures.     

Caspase involvement in the mitochondrial membrane depolarization during ganglioside-induced apoptosis of thymocytes

Ganglioside-induced apoptosis in mouse thymocytes was shown to be caspase-dependent, mitochondria being involved in the apoptosis-signaling pathway of GM1-, GD3-and GT1b-stimulated cells. According to their role in caspase-8-induced signaling cascades in thymocytes, these gangliosides can be divided into two groups, viz., those activating cell apoptosis by a mitochondrial route without the involvement of death receptors and caspase-8 (the so-called mitochondrial signaling cascade) (GD3), and those activating this process by receptor-mediated and mitochondrial routes (GM1 and GT1b). Anti-Fas antibodies that activate apoptosis of thymocytes by receptor pathway were used as a reference system. Cytofluorimetric studies of chromosomal DNA fragmentation revealed that effector caspase-3 is involved in apoptotic signaling cascades triggered by all the gangliosides under study. At the same time, the caspase-3 inhibitor Z-DEVD-FMK abolished the ganglioside-and antibody-induced depolarization of thymocyte mitochondrial membranes by a receptor-dependent route either partly (GM1 and GT1b) or completely (anti-Fas antibodies). Thymocytes stimulated by GD3 by a mitochondrial apoptotic route were an exception. Possible mechanisms of the caspase-3 involvement in the regulation of the activity of mitochondrial apoptosis-induced channels (MAC) are discussed and in particular, the role of proapoptotic proteins Bax/Bid.     

Complete nucleotide sequences of bovine alpha S2- and beta-casein cDNAs: comparisons with related sequences in other species

The nucleotide sequences corresponding to bovine alpha S2- and beta- casein mRNAs have been determined by cDNA analysis. Both sequences appear to be complete at their 5' ends. The nucleotide sequence of alpha S2-casein, when compared with the corresponding cavine A sequence, helps to define the boundaries of a large amino acid repeat (approximately 80 residues) whereas comparisons with the nucleotide sequences of rat gamma- and mouse epsilon-casein mRNAs also reveal extensive sequence similarities. An alignment of these four sequences shows that the divergence of their translated regions has been characterized by the duplication and deletion of discrete segments of sequence that probably correspond to exons. A high degree of nucleotide substitution is also found when the four sequences are compared, except for well-conserved leader-peptide and phosphorylation-site sequences and, to a lesser extent, the 5'-untranslated regions. Similar comparison of the bovine and rat beta-caseins shows that their divergence has involved a high rate of nucleotide substitution but that no major insertions or deletions of sequence have occurred. The several splice sites that have veen defined in the rat beta-casein gene are likely to have been conserved in the bovine. The contrasting evolutionary histories of the alpha- and beta-casein coding sequences correlate with the distinctive functions of these proteins in the casein micelle system in milk.      

Rapid ex vivo expansion of highly enriched human invariant natural killer T cells via single antigenic stimulation for cell therapy to prevent graft-versus-host disease

Background aims

CD1d-restricted invariant natural killer (iNK) T cells are rare regulatory T cells that may contribute to the immune-regulation in allogeneic stem cell transplantation (ASCT). Here, we sought to develop an effective strategy to expand human iNK T cells for use in cell therapy to prevent graft-versus-host disease (GVHD) in ASCT.

A biologically active fragment of the differentiation factor of the HL-60 line cells: Identification and properties

Six-membered peptide fragment TGENHR (HLDF-6) was identified in the HL-60 cell culture of human promyelocyte leukemia treated with retinoic acid when studying the differentiation factor HLDF of this cell line. HLDF-6 retains the ability of the full-size factor to induce the differentiation and arrest the proliferation of the starting HL-60 cells. It was shown that the synthetic peptide HLDF-6 has no specific receptors on the surface of the HL-60 cells but can affect the binding of interleukin IL-1β, a cytokine involved in proliferation, to the cell surface. It was found on a model of transplantable NSO myeloma that HLDF-6 has an antitumor activity.     

Chitosan nanoparticles targeted to the tumor-associated ganglioside GD2

Methodological approaches to the creation of nanoparticles based on chitosan derivatives and targeted to the GD2-positive tumor cells were developed. The GD2-specific monoclonal antibodies and their Fab-fragments and scFv-fragments were obtained and studied as vector molecules. Various methods for covalent conjugation of these molecules to the nanoparticles were also studied. It was shown that site-specific conjugation of scFv-fragments of GD2-specific antibodies to the chitosan nanoparticles by using a reagent BMPS is the optimal approach to create targeted chitosan-based nanoparticles directed to tumor-associated ganglioside GD2.     

The nuclease activity of the endogenous differentiation factor of the HL-60 cell line

A structural homology between the endogenous differentiation factor of the HL-60 cell line of promyelocyte leukemia (HLDF) and several DNA/RNA-binding and DNA/RNA-hydrolyzing proteins was revealed, and expression of thehldf gene in prokaryotic systems was studied. On the basis of these experiments, the amino acid sequence of an 8-membered fragment of HLDF with potential nuclease activity was identified. The synthetic octapeptide RRWHRLKE was shown to be capable of the cleavage of RNA, linear DNA from phage λ, and all forms of plasmid DNA. We established that treatment of the HL-60 cell culture with this peptide (10⁻⁶ M) results in an increase in the number of apoptotic cells and suggested that HLDF is involved in processes of apoptosis.     

Biologically relevant effects of mRNA amplification on gene expression profiles

Background  

Gene expression microarray technology permits the analysis of global gene expression profiles. The amount of sample needed limits the use of small excision biopsies and/or needle biopsies from human or animal tissues. Linear amplification techniques have been developed to increase the amount of sample derived cDNA. These amplified samples can be hybridised on microarrays. However, little information is available whether microarrays based on amplified and unamplified material yield comparable results.