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

Prirodno osvetljenje u adaptiranim industrijskim objektima

This paper assesses the potential of historical industrial buildings to be reused as office spaces. Belgrade’s industrial heritage has been classified according to the criteria that influence visual comfort, including glazing area, floor depth, and orientation. Daylight performance of two representative buildings has been analyzed using daylight factor, point in time illuminance and spatial daylight autonomy. Potential improvement strategies that would not have a negative impact on the historical character of buildings have then been discussed. Further studies include increased internal surface reflectance and introduction of roof-lights. The impact of roof-lights on the annual cooling and heating load has been addressed in parallel. Since LEED is the dominant sustainability assessment tool in Serbia, preliminary compliance with LEED v4 Daylight credit has been assessed for all options. The methodology and findings can be applied to a wide range of industrial buildings in similar climatic conditions.U radu je analizirana prenamena industrijskih objekata izgrađenih pre Drugog svetskog rata u poslovne. Industrijsko nasleđe Beograda je klasifikovano prema kriterijuma koji utiču na vizuelni komfor, uključujući veličinu prozora, dubinu osnove i orijentaciju objekata. Količina prirodne svetlosti je analizirana korišćenjem faktora dnevne osvetljenosti (daylight factor), nivoa osvetljenosti (point in time illuminance) i prostorne autonomije dnevne svetlosti (spatial daylight autonomy). Nakon analiza, razmatrane su potencijalne strategije za unapređenje nivoa osvetljenosti koje ne bi imale negativan uticaj na istorijski karakter zgrada. Dalje analize uključuju povećanje vrednosti refleksije površina prostorije i uvođenje krovnih prozora. Uticaj krovnih prozora na godišnju potrebnu energiju za grejanje i hlađenje objekata je takođe razmatrano. LEED je dominantan program za sertifikaciju poslovnih zgrada u Srbiji. Stoga, rad uključuje preliminarane analize svih opcija za usklađenost sa zahtevima LEED v4 kredita “Prirodno osvetljenje”. Metodologija i zaključci ovog istraživanja mogu se primeniti na širok spektar industrijskih objekata u sličnim klimatskim uslovima

Effect of different growth promoters on broiler performance and gut morphology

A total of 240 Hybro broilers was divided into 4 groups. These groups were fed a complete corn/soybean based diet with and without addition of antibiotic growth promoters (AGP, Flavomycin® 15 ppm, Intervet), direct feed microbials (DFM, All-Lac® 1kg/T, Alltech Inc. USA) and mannanoligosaccharide (MOS) (Bio-MOS® 2kg/T, Alltech Inc. USA). Chickens were introduced into the experiment after hatching. At day 42 of trial, all broilers were conventionaly sacrificed in a slaughter plant and slaughter performances were measured. Samples of intestines with its content from 6 average broilers (randomly selected) from each group (n=24) were taken for examination. At the end of the trial, body weight (BW) and body weight gain (BWG) of broilers fed the diet containing Bio-MOS® (1915.23 and 44.58 g), AGP (1869.40 and 43.50 g) and DFM (1855.50 and 43.17g) were significantly higher than in birds of the control group (1815.67 and 41.96 g). When compared with the control group (91.19 g), ADFI (average daily feed intake) was also significantly reduced when Bio-MOS® (81.84 g), DFM (83.50g) or AGP (86.16 g) were supplemented which lead to a lower feed conversion ratio (FCR), 2.17 vs. 1.83, 1.93 and 1.98 kg, respectively. A significantly lower pH of the intestinal content of intestines (duodenum, ileum, caecum) was observed in groups fed DFM (6.16, 6.46 and 6.72) and Bio-MOS® (6.25, 6.50 and 6.78), compared with the control (6.55, 6.81 and 7.21) and AGP (6.61, 6.87 and 7.14) group. The use of DFM and Bio-MOS® increased length and width of intestinal villa and decreased depth of crypts, while the number of goblet cells did not significantly differ among experimental groups. In conclusion, Bio-MOS® and DFM exhibited nutritional, pharmacological and economic advantages over antibiotic growth promoters

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

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

Rendgenska, ultrazvučna i endoskopska dijagnostika određenih oboljenja želuca kod pasa

In order to broaden the range of diagnostic methods for determining stomach diseases and to make these methods a greater part of the daily clinical practice, the procedures of radiological, ultrasound and endoscopic examinations should be worked out in more detail, their limits and possibilities should be established, the topographic-anatomical and morphological status of the stomach should be described in a radiological, ultrasound and endoscopic picture, and the most represented stomach diseases should be defined as well. The paper presents the results of six-month investigations of stomach diseases in dogs of different breeds and ages. A total of 20 animals were examined: seven German Shepherds, four golden retrievers, three rottweilers, two dobermans, two big schauzers, and two mixed-breed dogs. Cases of acute gastritis, chronic ulcerous gastritis, foreign body cases, and stomach tumors have been described.Da bi se spektar dijagnostičkih metoda oboljenja želuca proširio i bio prisutniji u svakodnevnoj kliničkoj praksi, poželjno je da se detaljnije razradi postupak radiološkog, ultrazvučnog i endoskopskog pregleda, utvrde njihove granice i mogućnosti, opiše topografsko-anatomski i morfološki status želuca u radiološkoj, ultrazvučnoj i endoskopskoj slici i da se utvrdi koja su to najzastupljenija oboljenja ovog organa. U radu su prikazani rezultati šetomesečnih ispitivanja oboljenja želuca na psima različitih rasa i uzrasta. Ukupno je pregledano dvadeset jedinki i to: sedam nemačkih ovčara, četiri zlatna retrivera, tri rotvajlera, dva dobermana, dva velika šnaucera i dva mešanca. Opisani su slučajevi akutnog gastritisa, hroničnog ulceroznog gastritisa, stranog tela i tumora želuca

Morfološke, imunofenotipske i kliničke karakteristike limfocitno - plazmocitnog duodenitisa pasa

The most important morphological feature of the immune response in lymphocytic-plasmacytic duodenitis (LPD) in dogs is the presence of a mononuclear infiltrate in the duodenal mucosa. The ethiopatogenesis of this disease is still unknown, nor are known all the immunophenotypic characteristics of the infiltrate cells, which would be of help in the elucidation of the pathogenesis of LPD. The study involved 60 adult dogs of different breeds and both sexes: 54 dogs with symptoms of diarrhea and vomiting that lasted longer than three weeks and six clinically healthy dogs that served as controls. Hematological and biochemical analysis of blood, radiology, ultrasound and endoscopic diagnosis were carried out. Samples of duodenal mucosal biopta were examined by histopathological and immunohistochemical methods. During endoscopic examination, duodenal wall thickening, mucosal edema and hyperemia were observed in 46 dogs. Microscopic studies showed on the epithelial cells of the duodenum degenerative changes, and sometimes desquamation. Most of the intestinal crypts were dilated, irregularly shaped and filled with detritus. On immunohistochemical examination of samples of the duodenum in a number of dogs with LPD a predominant expression of CD3 + T lymphocytes was observed.Najvažnija morfološka karakteristika imunskog odgovora kod limfocitnoplazmocitnog duodenitisa (LPD) pasa je nalaz infiltrata mononuklearnih ćelija krvi u sluzokoži duodenuma. Do danas etiopatogeneza ove bolesti pasa nije u svemu poznata, a nisu poznate ni sve imunofenotipske karakteristike ćelija u infiltratu, koje bi bile značajne za razumevanje etiopatogeneze LPD. Ispitivanjem je bilo obuhvaćeno ukupno 60 odraslih pasa, različitih rasa i oba pola: 54 psa sa simptomima dijareje i povraćanja koje je trajalo duže od tri nedelje i šest klinički zdravih pasa koji su služili kao kontrola. Urađene su hematološke i biohemijske analize krvi, a zatim je sprovedena radiološka, ultrazvučna i endoskopska dijagnostika. Uzorci mukoze duodenuma, uzeti biopsijom, ispitani su histopatološkim i imunohistohemijskim metodama. Pri endoskopskom pregledu, kod 46 pasa je zapažen zadebljali zid duodenuma, hiperemičnost i edem sluznice. Mikroskopskim ispitivanjima, na epitelnim ćelijama duodenuma uočene su degenerativne promene, a nekada i deskvamacija. Većina crevnih kripti je bila dilatirana, nepravilnog oblika i ispunjena detritusom. Imunohistohemijskim ispitivanjem uzoraka duodenuma kod većine pasa sa LPD uočena je dominacija CD3+T limfocita

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

Involvement of ubiquitous and tale transcription factors, as well as liganded RXRα, in the regulation of human SOX2 gene expression in the NT2/D1 embryonal carcinoma cell line

SOX2 transkripcioni faktor ima ključnu ulogu u procesima embrionalnog razvića i predstavlja univerzalni marker pluripotentnih matičnih ćelija. S obzirom na funkcionalnu redundantnost i preklapajući profil ekspresije članova SOXB1 podgrupe tokom razvića, cilj ovog rada bio je da ispita potencijalne zajedničke aspekte regulacije ekspresije SOX2 i SOX3 gena. Naime, ispitivan je uticaj odabranih transkripcionih faktora na regulaciju ekspresije SOX2 gena u NT2/D1 ćelijskoj liniji. Analizirani su oni faktori za koje je prethodno pokazano da su uključeni u modulaciju aktivnosti humanog SOX3 gena. Rezultati ovih istraživanja ukazuju da opšti transkripcioni faktori (NF-Y, Sp1 i MAZ), članovi TALE familije proteina (Pbx1 i Meis1), kao i retinoičnom kiselinom aktiviran nuklearni receptor RXRα dovode do povećane ekspresije SOX2 proteina. Ispitivanje transkripcionih faktora uključenih u regulaciju ekspresije SOX gena je značajno za bolje razumevanje signalnih puteva koji su aktivni u pluripotentnim matičnim ćelijama.SOX2 is a key transcription factor in embryonic development representing a universal marker of pluripotent stem cells. Based on the functional redundancy and overlapping expression patterns of SOXB1 subgroup members during development, the goal of this study has been to analyze if some aspects of regulation of expression are preserved between human SOX2 and SOX3 genes. Thus, we have tested several transcription factors previously demonstrated to play roles in controlling SOX3 gene activity for potential participation in the regulation of SOX2 gene expression in NT2/D1 cells. Here we report on the activation of SOX2 expression by ubiquitous transcription factors (NF-Y, Sp1 and MAZ), TALE family members (Pbx1 and Meis1), as well as liganded RXRα. Elucidating components involved in the regulation of SOX gene expression represent a valuable contribution in unraveling the regulatory networks operating in pluripotent embryonic cells

Nutritive value of green mass and hay from the Divčibare region

With the aim of forming a complete observation of the botanical composition and nutritive value of pasture and meadow hay from the Divčibare region, which is a typically hilly-mountain area in Serbia, investigations were designed to evaluate the quality of pasture and hay (chemical composition and energy value), as well as the losses which are results of drying pasture. Twelve samples of grass were taken from three different altitudes and after mowing, samples of hay were also taken from the same meadow. In a conventional chemical analysis (Weende) the chemical composition of grass and hay were determined. On the basic of this analysis nutritive values were calculated. Pasture and hay from the Divčibare region have a satisfying chemical composition, they can be used as a forage base and a source of nutrients for ruminants, predominantly, lowproducing livestock. The difference in content of proteins among samples from some localities might be bound with the content of fabaceae. The increase fibre content with the proportional decrease content of protein had a negative influence on the nutritive value in the samples of grass and hay. The Net Energy Lactation (NEL) of hay from the investigated region is near the lower limit reported as average for pasture and meadow hay. Significant higher nutritive values of green mass, calculated on dry matter (NEL 8.35 MJ/kg, SV 0.78 SJ/kg) when compared to the nutritive value of hay (NEL 8.12 MJ/kg, SV 0.74 SJ/kg) show that losses are results of haymaking. Nutrients losses during drying are in correlation with losses of proteins, lipids and mineral matter, but there is an increase in fiber. The change in chemical composition due to the described losses during drying and the negative influence on organic matter digestibility, led to a reduction in the energy value of the investigated samples.Со цел да формираме комплетна слика за ботаничкиот состав и хранливата вредност на пашата и ливадското сено од регионот на Дивчибаре, кој е типично високопланинско подрачје во Србија, извршивме евалуација на квалитетот на пашата и сеното (хемиски состав и енергетска вредност) и на загубите кои се резултат на сушење на пашата. Беа земeни дванаесет проби од трева од три различни височини, и тоа по цутењето, додека пробите од сено беа земани секогаш од истото место. Со конвенционална хемиска анализа (методот на Weende) беше одреден хемискиот состав на тревата и сеното. Врз база на тие анализи беше пресметана хранливата вредност. Пашата и сеното од регионот на Дивчибаре имаа задоволувачки хемиски состав. Тие можат да бидат користени како крмна база и извор на хранливи материи за преживари, предоминатно, и за послабо продуктивни животни. Разликите во содржината на протеини кај повеќето проби од различни локалитети можеби се поврзани со присуството на растенија од фамилијата Fabaceae. Зголемувањето на содржината на влакно со пропорционалното намалување на содржината на протеини има негативно влијание на хранливата вредност на пробите од трева и сено. Нето-енергијата за лактација (NEL) на сеното од истражуваниот регион е блиску до намалениот лимит рефериран во литературата како просек за пашата и ливадското сено. Утврдена е значително повисока хранлива вредност на зелената маса, калкулирана на сува материја (NEL 8.35 MJ/kg; SV 0.78 SJ/kg), во споредба со хранливата вредност на сеното (NEL 8.12 MJ/kg, SV 0.74 SJ/kg), што покажува дека загубите се резултат на обработката на сеното. Загубите во хранливи материи за време на сушењето се во корелација со загубите на протеини, липиди и минерални материи, но не со зголемувањето на влакното. Промените во хемискиот состав опишани како загуби во текот на сушењето и негативното влијание врз сварливоста на органските материи доведуваат до намалување на енергетската вредност на испитуваните проби