The Administration of Semax and HLDF-6 Peptides to Rats Regulates Protein Synthesis Rhythm in Hepatocytes and Corrects Senescent Disturbances

Russian Journal of Developmental Biology - To clarify the organizing effect of Semax and HLDF-6 peptides on the kinetics of protein synthesis in hepatocytes, in addition to an in vitro study...     

EST-based gene discovery in pig: virtual expression patterns and comparative mapping to human

A molecular understanding of porcine reproduction is of biological interest and economic importance. Our Midwest Consortium has produced cDNA libraries containing the majority of genes expressed in major female reproductive tissues, and we have deposited into public databases 21,499 expressed sequence tag (EST) gene sequences from the 3 end of clones from these libraries. These sequences represent 10,574 different genes, based on sequence comparison among these data, and comparison with existing porcine ESTs and genes indicate as many as 4652 of these EST clusters are novel. In silico analysis identified sequences that are expressed in specific pig tissues or organs and confirmed the broad expression in pig for many genes ubiquitously expressed in human tissues. Furthermore, we have developed computer software to identify sequence similarity of these pig genes with their human counterparts, and to extract the mapping information of these human homologues from genome databases. We demonstrate the utility of this software for comparative mapping by localizing 61 genes on the porcine physical map for Chromosomes (Chrs) 5, 10, and 14. The following Accession numbers were assigned to our deposited sequences: BF701840 – BF704551, BF708383, BF708386 – BF713604, BG322266 – BG322271, BI398567 – BI405235, BQ597354 – BQ605166.     

Onset of rosette formation during spontaneous neural differentiation of hESC and hiPSC colonies

In vitro neural differentiation of human embryonic stem cells (hESCs) is an advantageous system for studying early neural development. The process of early neural differentiation in hESCs begins by initiation of primitive neuroectoderm, which is manifested by rosette formation, with consecutive differentiation into neural progenitors and early glial-like cells. In this study, we examined the involvement of early neural markers – OTX2, PAX6, Sox1, Nestin, NR2F1, NR2F2, and IRX2 – in the onset of rosette formation, during spontaneous neural differentiation of hESC and human induced pluripotent stem cell (hiPSC) colonies. This is in contrast to the conventional way of studying rosette formation, which involves induction of neuronal differentiation and the utilization of embryoid bodies. Here we show that OTX2 is highly expressed at the onset of rosette formation, when rosettes comprise no more than 3–5 cells, and that its expression precedes that of established markers of early neuronal differentiation. Importantly, the rise of OTX2 expression in these cells coincides with the down-regulation of the pluripotency marker OCT4. Lastly, we show that cells derived from rosettes that emerge during spontaneous differentiation of hESCs or hiPSCs are capable of differentiating into dopaminergic neurons in vitro, and into mature-appearing pyramidal and serotonergic neurons weeks after being injected into the motor cortex of NOD-SCID mice.     

A new crystal form of the complex between seryl-tRNA synthetase and tRNA(Ser) from Thermus thermophilus that diffracts to 2.8 A resolution.

Two distinct complexes between seryl-tRNA synthetase and tRNA(Ser) from Thermus thermophilus have been crystallized using ammonium sulphate as a precipitant. Form III crystals grow from solutions containing a 1:2.5 stoichiometry of synthetase dimer to tRNA. They are of monoclinic space group C2 with unit cell dimensions a = 211.6 A, b = 126.8 A, c = 197.1 A, beta = 132.4 degrees and diffract to about 3.5 A. Preliminary crystallographic results show that the crystallographic asymmetric unit contains two synthetase dimers. Form IV crystals grow from solutions containing a 1:1.5 stoichiometry of synthetase dimer to tRNA. They are of orthorhombic space group P2(1)2(1)2(1) with unit cell dimensions a = 124.5 A, b = 128.9 A, c = 121.2 A and diffract to 2.8 A resolution. Preliminary crystallographic results show that these crystals contain only one tRNA molecule bound to a synthetase dimer.     

Chromosomal and regional localization of the loci for IGKC,IGGC, ALDB,HOXB, GPT,and PRNP in the American mink (Mustela vison): comparisons with human and mouse

Chromosomal localization of the genes for gamma- and kappa-immunoglobulins (IGGC and IGKC, respectively), aldolase B (ALDB), prion protein (PRNP), homeo box B (HOXB), and glutamate pyruvate transaminase (GPT) were determined with the use of mink-rodent hybrid cells. Analysis of segregation of the mink markers and chromosomes in these hybrid cells allowed us to assign the gene for HOXE to Chromosome (Chr) 8, IGGC to Chr 10, PRNP and IGKC to Chr 11, ALDB to Chr 12, and GPT to Chr 14 in mink. Furthermore, using a set of mink-mouse hybrid cells carrying fragments of mink Chr 8 of different sizes, we assigned the gene for HOXB to the pter-p26 region of the short arm of Chr 8. Comparative mapping of the genes of mink, human, and mouse, as well as other mammalian species, demonstrated that the mink genes HOXB, PRNP, ALDB, and IGGC are members of a conserved region shared by many mammalian species in common; the IGKC gene is a member of a conserved region common to carnivores and primates, not rodents; the GPT gene is a member of a syntenic gene group probably unique to the Mustelidae family or carnivores.     

Cancer hallmarks in induced pluripotent cells: New insights

Studies are beginning to emerge that demonstrate intriguing differences between human‐induced pluripotent stem cells (hiPSCs) and human embryonic stem cells (hESCs). Here, we investigated the expression of key members of the Nodal embryonic signaling pathway, critical to the maintenance of pluripotency in hESCs. Western blot and real‐time RT‐PCR analyses reveal slightly lower levels of Nodal (a TGF‐β family member) and Cripto‐1 (Nodal's co‐receptor) and a dramatic decrease in Lefty (Nodal's inhibitor and TGF‐β family member) in hiPSCs compared with hESCs. The noteworthy drop in hiPSC's Lefty expression correlated with an increase in the methylation of Lefty B CpG island. Based on these findings, we addressed a more fundamental question related to the consequences of epigenetically reprogramming hiPSCs, especially with respect to maintaining a stable ESC phenotype. A global comparative analysis of 365 microRNAs (miRs) in two hiPSC versus four hESC lines ultimately identified 10 highly expressed miRs in hiPCSs with >10‐fold difference, which have been shown to be cancer related. These data demonstrate cancer hallmarks expressed by hiPSCs, which will require further assessment for their impact on future therapies. J. Cell. Physiol. 225: 390–393, 2010. © 2010 Wiley‐Liss, Inc.     

Small cooperative activity of old rat hepatocytes may depend on composition of the intercellular medium

Ultradian oscillations of protein synthesis were used as a marker of hepatocyte synchronous cooperative activity producing a common rhythm in vitro; amplitude of the rhythm defines expression of the cell cooperation. Dense synchronous and sparse non-synchronous rat hepatocyte cultures on slides in a serum-free incubation medium 199 supplemented with 0.2 mg/ml albumin and 0.5 microg/ml insulin have been studied. The amplitude of the rhythm averaged approximately 2x in dense cultures of young (3 month old) rats than in old (2 year old) rats. But some cultures of young rats had the amplitude patterns similar to cultures of old rats, and vice versa. Addition to the medium of either 0.3 microM bovine brain gangliosides or 2 microM phenylephrine resulted in increase of the oscillation amplitude in dense cultures of old rats to the level inherent in young ones. Addition to the medium of 10% rat blood serum in non-synchronous sparse cultures from young rats resulted in detection of a protein synthetic rhythm. Although after serum from young rats, the rhythm expression was high, the rhythm after serum from old rats had been given was weak. Addition of gangliosides to old-rat serum resulted in synchronization of sparse cultures with amplitudes inherent of young-rat serum. The data tend to the conclusion that cell cooperation depends to a greater extent on the composition of the medium rather than on the age of the cell or animal.     

A Mouse Model of Human Primitive Neuroectodermal Tumors Resulting from Microenvironmentally-Driven Malignant Transformation of Orthotopically Transplanted Radial Glial Cells

There is growing evidence and a consensus in the field that most pediatric brain tumors originate from stem cells, of which radial glial cells constitute a subtype. Here we show that orthotopic transplantation of human radial glial (RG) cells to the subventricular zone of the 3rd ventricle - but not to other transplantation sites - of the brain in immunocompromised NOD-SCID mice, gives rise to tumors that have the hallmarks of CNS primitive neuroectodermal tumors (PNETs). The resulting mouse model strikingly recapitulates the phenotype of PNETs. Importantly, the observed tumorigenic transformation was accompanied by aspects of an epithelial to mesenchymal transition (EMT)-like process. It is also noteworthy that the tumors are highly invasive, and that they effectively recruit mouse endothelial cells for angiogenesis. These results are significant for several reasons. First, they show that malignant transformation of radial glial cells can occur in the absence of specific mutations or inherited genomic alterations. Second, they demonstrate that the same radial glial cells may either give rise to brain tumors or differentiate normally depending upon the microenvironment of the specific region of the brain to which the cells are transplanted. In addition to providing a prospect for drug screening and development of new therapeutic strategies, the resulting mouse model of PNETs offers an unprecedented opportunity to identify the cancer driving molecular alterations and the microenvironmental factors that are responsible for committing otherwise normal radial glial cells to a malignant phenotype.     

Sensitivity of starfish oocytes and whole, half and enucleated embryos to cytotoxic neuropharmacological drugs

In early embryos of eight starfish species both usual sensitivity and supersensitivity to cytotoxic neurochemicals have been found. This sensitivity is unaffected by the removal of the cell nucleus. In the starfish Aphelasterias japonica the supersensitivity in the embryos' red halves is higher and in embryos' white halves lower than in the whole embryos.     

Single short-term signal that enhances cooperative activity of old rat hepatocytes acts for several days

Primary cultures of rat hepatocytes were studied in serum-free medium. Ultradian protein synthesis rhythm was used as a marker of overall cell synchronization and cooperation amongst the population. The level of synchronization was determined by amplitudes of the rhythm. Low synchronization of old rat hepatocytes can be enhanced by addition of either gangliosides or phenylephrine to the medium. Incubation of cultures with gangliosides lasted for 2.5 h, while action of phenylephrine was only for 2 min. The amplitude of protein synthesis rhythm was increased 1.5-2 times. In cultures transferred to a fresh normal medium, this increased amplitude was observed for at least 2-3 days. Thus, both gangliosides and phenyleprine are triggers, which, as shown earlier, initiated calcium-dependent processes in the cytoplasm. The results are discussed in the light of concept of the cell self-organization by a direct cell-cell communication.