Establishment and initial characterization of SOX2-overexpressing NT2/D1 cell clones

SOX2, a universal marker of pluripotent stem cells, is a transcription factor that helps control embryonic development in vertebrates; its expression persists in neural stem/progenitor cells into adulthood. Considering the critical role of the SOX2 transcription factor in the regulation of genes required for self-renewal and pluripotency of stem cells, we developed and characterized SOX2-overexpressing NT2/D1 cell clones. Using Southern blot and semi-quantitative RT-PCR, we confirmed integration and expression of exogenous SOX2 in three NT2/D1 cell clones. Overexpression of the SOX2 gene was detected in two of these clones. SOX2 overexpression in NT2/D1 cell clones resulted in altered expression of key pluripotency genes OCT4 and NANOG. Furthermore, SOX2-overexpressing NT2/D1 cell clones entered into retinoic acid-dependent neural differentiation, even when there was elevated SOX2 expression. After 21 days of induction by retinoic acid, expression of neural markers (neuroD1 and synaptophysin) was higher in induced cell clones than in induced parental cells. The cell clone with SOX2 overexpression had an approximately 1.3-fold higher growth rate compared to parental cells. SOX2 overexpression did not increase the population of cells undergoing apoptosis. Taken together, we developed two SOX2-overexpressing cell clones, with constitutive SOX2 expression after three weeks of retinoic acid treatment. SOX2 overexpression resulted in altered expression of pluripotency-related genes, increased proliferation, and altered expression of neural markers after three weeks of retinoic acid treatment

An exploratory study of heavy domain wall fermions on the lattice

We report on an exploratory study of domain wall fermions (DWF) as a lattice regularisation for heavy quarks. Within the framework of quenched QCD with the tree-level improved Symanzik gauge action we identify the DWF parameters which minimise discretisation effects. We find the corresponding effective 4$d$ overlap operator to be exponentially local, independent of the quark mass. We determine a maximum bare heavy quark mass of $am_h\approx 0.4$, below which the approximate chiral symmetry and O(a)-improvement of DWF are sustained. This threshold appears to be largely independent of the lattice spacing. Based on these findings, we carried out a detailed scaling study for the heavy-strange meson dispersion relation and decay constant on four ensembles with lattice spacings in the range $2.0-5.7\,\mathrm{GeV}$. We observe very mild $a^2$ scaling towards the continuum limit. Our findings establish a sound basis for heavy DWF in dynamical simulations of lattice QCD with relevance to Standard Model phenomenology.Comment: 23 pages, 8 figure

PCR amplification and sequence analysis of the rat Sox3 gene

Sox3 gen je jedan od markera najranijih faza razvića nervnog sistema kičmenjaka koji je uključen u kontrolu diferencijacije nervnih prekursora. Uprkos činjenici da je genom pacova sekvenciran i javno dostupan, samo parcijalna sekvenca Sox3 gena ove vrste je bila deponovana u bazi podataka. U ovom radu smo primenom PCR-a, sekvenciranja i bioinformatičke analize generisali kompletnu kodirajuću sekvencu Sox3 gena pacova. Analiza dobijene sekvence je pokazala da Sox3 gen kodira protein od 449 amino kiselina. Uporedna analiza ortologih SOX3 proteina pacova i čoveka pokazala je visok stepen evolutivne očuvanosti. Identifikacija i karakterizacija Sox3 gena pacova doprineće boljem razumevanju njegove uloge tokom razvića nervnog sistema i omogućiće bolji uvid u evoluciju ovog gena kod vertebrata.The Sox3 gene is considered to be one of the earliest neural markers in vertebrates, playing a role in specifying neuronal fate. Despite the completion of a rat genome sequencing project, only a partial sequence of the rat Sox3 gene has been available in the public database. Using PCR, sequencing, and bioinformatics tools, in this study we have determined the complete coding sequence of the rat Sox3 gene encoding 449 amino acids. Comparative analysis of rat and human SOX3 proteins revealed a high degree of conservation. Identification of the rat Sox3 gene sequence would help in understanding the biological roles of this gene and provide insight into evolutionary relationships with vertebrate orthologs

3D random fiber composites as a repair material for damaged honeycomb cores

Povećana upotreba kompozitnih materijala donela potrebu za razvojem pouzdanih i efikasnih tehnika popravki. Kao i sve druge noseće metalne strukture i kompozitne strukture sklone su oštećenjima. Razvijene tehnike popravki metalnih konstrukcije nisu direktno primenljive na kompozite i problem popravki kompozitnih struktura mora biti detaljno proučen. U ovom radu analizirana je primena kompozitnih materijala sa slučajnim rasporedom vlakana, kao materijalom za popravku oštećenih saćastih jezgara visoke gustine sendvič konstrukcija. Pomoću Panove teorije izveden je izraz potrebnog zapreminskog udela vlakana materijala popravke i određeno je kompletno naponsko deformaciono stanje popravljene kompozitne stukture metodom konačnih elemenata. Zaključeno je da kompozitni materijali sa slučajnim rasporedom vlakana predstavljaju veoma dobre kandidate za popravku oštećenih kompozitnih struktura sa saćastim jezgrima.The increased use of composites has rendered the need for development of reliable and efficient repair techniques. Like all other metallic structures composite structures are prone to variety of damage. Repair techniques developed for metallic structures are not directly applicable to composites and problem of composite repair has to be investigated in great detail. In the present paper application of 3D random fiber polymer composites, as a repair material for damaged high density honeycomb cores in sandwich structures is investigated. Based on Pan 's theory the expression for matching fiber volume fraction of the repair material is derived and complete stress-strain field, of the repaired sandwich structure is determined using finite element approach. It is concluded that the 3D random fiber composites represent very good candidates as repair materials for damaged honeycomb structured composites

Electrical characteristics of female and male human skin

Bioimpedance spectroscopy (BIS) is a popular method for characterizing the electrical properties of biological tissues. In this study, BIS measurement data of female and male human skin were analyzed and compared. The electrical characteristics of tissue were followed according to four-parameters of the Cole-Cole model: low frequency resistance R0; high frequency resistance R∞; relaxation time t and parameter a. Individual electrical characteristics of human skin were determined for 30 women and 30 men. The distribution and one-way analysis of variance (one-way ANOVA) of the Cole-Cole parameters R0, R∞, t, a within the human population indicated their different dependence on gender. Parameter a, which is higher in the female subjects (a =0.83±0.03) than in the male subjects (a=0.7±0.05), is strongly dependent on gender (p=0). Parameter R∞ also significantly depends on gender (p=0.002), while t and R0 seem to be slightly related to gender (p>0.05). [Acknowledgments - This work was supported by the Ministry of Education and Science of the Republic of Serbia (41006).

Stress effects on the phosphorylation of c-jun-nterminal kinases and on nuclear translocation of hsp70 in rat hippocampus

Glucocorticoids have diverse effects in cellular processes in hippocampus (HIPPO) under stress. Beside genomic pathways, their effects are also mediated by direct activation of subfamily of mitogen-activated protein kinases termed, c-Jun-Nterminal kinases (JNKs). We analysed the phosphorylation status of cytoplasmic and nuclear JNK isoforms, and expression of its inhibitor Hsp70 protein in HIPPO of rats exposed to diverse types of stress. Activity of JNK1 in cytoplasm and nucleus was decreased in all types of stress, while the activity of cytoplasmic JNK2/3 was markedly higher in acute stress, and unaltered or lowered in chronic and combined stress. Hsp70 was significantly decreased in cytoplasm and increased in nucleus under all stress conditions indicating its cytoplasmic-nuclear translocation.Physical chemistry 2008 : 9th international conference on fundamental and applied aspects of physical chemistry; Belgrade (Serbia); 24-28 September 200

Members of the CREB/ATF and AP1 family of transcription factors are involved in the regulation of SOX18 gene expression

The SOX18 transcription factor plays an important role in endothelial cell specification, angiogenesis and atherogenesis. By profiling transcription factor interactions (TranSignal TM TF Protein Array) we identified several transcription factors implicated in angiogenesis that have the ability to bind to the SOX18 optimal promoter region in vitro. In this report we focused our attention on distinct transcription factors identified by the array as belonging to AP-1 and CREB/ATF protein families. In particular, we analyzed the effects of CREB, JunB, c-Jun and ATF3 on SOX18 gene expression. Functional analysis revealed that CREB acts as a repressor, while JunB, c-Jun and ATF3 act as activators of SOX18 promoter activity. Our findings indicate that a transcriptional network that includes CREB, JunB, c-Jun and ATF3 could be involved in angiogenesis-related transcriptional regulation of the SOX18 gene

Determination of the sulphur balance in coal during combustion in small and large fuel beds

U toku ovih ispitivanja eksperimentalno smo provjeravali uobičajeni proračun emisije sumpora na osnovu tzv. »sagorivog sumpora« u uglju. Mjerenja sadržaja sumpora u dimnim plinovima su pokazala da se ovakvim proračunima, koji se baziraju na određivanju tzv. »vezanog sumpora« u laboratorijskim uslovima spaljivanjem goriva do 800°C, može načiniti greška i do 100%. Emisija sumpora iz velikih ložišta za centralno grijanje kreće se otprilike 90%, dok je pretpostavljena emisija iz podataka o »sagorivom sumporu« trebala da iznosi otprilike 50%. Emisije sumpora iz sobnih peći kretale su se od 56 do 88% zavisno od konstrukcije, te od načina loženja i režima rada peći. I u ovom su slučaju emisije redovno bile više od očekivanih emisija »sagorivog sumpora«. Rezu!tati ovih mjerenja nam pokazuju da se prilikom projektovanja i proračuna emisije i visine dimnjaka ne možemo osloniti samo na podatke o sadržaju »sagorivog sumpora« u gorivu nego na eksperimentalne podatke o bilansu sumpora, koji se sve češće mogu naći u stranoj tehničkoj literaturi.The usual rough estimate of sulphur emission was checked experimentally on the basis of the sulphur content in coal known as »burnt sulphur«. The measurements of the content of sulphur in stack gases proved that such an estimate based on the measuring of sulphur in ash known as »bound sulphur« under laboratory conditions of burning fuel up to 800°C, can result in an error of up to 100%. The emission of sulphur from solid fuel beds used for central heating comes to 90% while the supposed emission based on »burnt sulphur« should be about 50%. The sulphur emission from domestic stoves reached 56% to 88% depending on stove design, coal burning and on operating conditions. The emissions were always greater than the expected emissions of »burnt sulphur«. The results of these measurements indicate that we cannot rely only on the information concerning »burnt sulphur« content when the design and calculation of chimney emission and height is carried out, but the experimental data on sulphur balance should also be taken into account. Such data can be more and more often found in foreign technical literature

Members of the CREB/ATF and AP1 family of transcription factors are involved in the regulation of SOX18 gene expression

The SOX18 transcription factor plays an important role in endothelial cell specification, angiogenesis and atherogenesis. By profiling transcription factor interactions (TranSignal TM TF Protein Array) we identified several transcription factors implicated in angiogenesis that have the ability to bind to the SOX18 optimal promoter region in vitro. In this report we focused our attention on distinct transcription factors identified by the array as belonging to AP-1 and CREB/ATF protein families. In particular, we analyzed the effects of CREB, JunB, c-Jun and ATF3 on SOX18 gene expression. Functional analysis revealed that CREB acts as a repressor, while JunB, c-Jun and ATF3 act as activators of SOX18 promoter activity. Our findings indicate that a transcriptional network that includes CREB, JunB, c-Jun and ATF3 could be involved in angiogenesis-related transcriptional regulation of the SOX18 gene

Regulation of the SOX3 Gene Expression by Retinoid Receptors

Sox3/SOX3 gene is considered to be one of the earliest neural markers in vertebrates. Despite the mounting evidence that Sox3/SOX3 is one of the key players in the development of the nervous system, limited data are available regarding the transcriptional regulation of its expression. This review is focused on the retinoic acid induced regulation of SOX3 gene expression, with particular emphasis on the involvement of retinoid receptors. Experiments with human embryonal carcinoma cells identified two response elements involved in retinoic acid/retinoid X receptor-dependent activation of the SOX3 gene expression: distal atypical retinoic acid-response element, consisting of two unique G-rich boxes separated by 49 bp, and proximal element comprising DR-3-like motif, composed of two imperfect hexameric half-sites. Importantly, the retinoic acid-induced SOX3 gene expression could be significantly down-regulated by a synthetic antagonist of retinoid receptors. This cell model provides a solid base for further studies on mechanism(s) underlying regulation of expression of SOX3 gene, which could improve the understanding of molecular signals that induce neurogenesis in the stem/progenitor cells both during development and in adulthood