Methylation Status of CYP27B1 and IGF2 Correlate to BMI SDS in Children with Obesity

OBJECTIVE: Worldwide increasing childhood obesity is due to interactions between environmental and genetic factors, linked together by epigenetic mechanisms such as DNA methylation. METHODS: 82 obese children (>95th BMI percentile , age: 3-18 years) were included. Anthropometric data, metabolic parameters, 25-OH vitamin D (25OHD), and pubertal status were recorded, 24-hour blood pressure monitoring was performed. BMI standard deviation score (SDS) was calculated. Using candidate gene approach, obesity- (insulin-like growth factor 2 (IGF2), proopiomelanocortin (POMC)) and vitamin D metabolism-related genes (1-alfa-hydroxylase (CYP27B1), VDR) regulated by DNA methylation were selected. After isolating DNA from peripheral blood, bisulfite conversion, bisulfite specific polymerase chain reaction (BS-PCR), and pyrosequencing were carried out. RESULTS: No significant correlation between 25-OHD and metabolic parameters and DNA methylation status, but a tendency of positive correlation between VDR methylation status and 25-OHD (r = 0.2053,p = 0.066) were observed. Significant positive correlations between BMI SDS and CYP27B1 hypermethylation (r = 0.2371,p = 0.0342) and a significant negative correlation between IGF2 hypomethylation and BMI SDS (r = -0.305,p = 0.0059) were found. Conclusions Rate of obesity shows correlation with DNA methylation. Hypomethylation of IGF2 and hypermethylation of CYP27B1 genes might positively influence the rate of BMI observed in obese children

Szerkezeti biológia = Structural biology

Kiroptikai spektroszkópia (ECD, VCD) - Az abszolút konfiguráció meghatározása - Gyűrűs és lineáris peptid modellek konformációjának meghatározása - Királis diródium komplexek ECD és VCD vizsgálata Fehérje NMR és modellezés - Peptidek számitástechnikai vizsgálata - Fehérjék NMR vizsgálata Fehérje röntgenkrisztallográfia - Egy prolil-oligopeptidáz (POP) komplexei kristályszerkezetének meghatározása - A kalmodulin (CaM) vizsgálata - A molekuláris felismerés szerkezeti vonatkozásai - A Pyrococcus horikoshii acilamino-peptidáz (AAP) kristályszerkezetének meghatározása Biokémiai vizsgálatok - A scallop-peptid vizsgálata három kristályközegben - A miozin-6 utolsó (tail) doménje szerkezetének predikciója - A coiled-coil és rendezetlen fehérje szegmensek kereszt-predikciójának analízise - Öt természetes és két szintetikus, töltéssel rendelkező ?-hélix lánc mikroszekundum időskálájú molekuladinamikai szimulációja vizes közegben - Az LC8 dynein centrális (hub) fehérje könnyű láncának (DYNLL) vizsgálata Immunológiai vizsgálatok - Az SHP-2 tirozin-foszfatáz vizsgálata. Különböző szerkezetű foszfopeptidek tanulmányozása - Egy biotinilezett kollagén epitóp peptid, az extravidin-peptid complex és extravidin szerkezetvizsgálata Közlemény: 36 | Chiroptical spectroscopy (ECD, VCD) - Determination of the absolute configuration - Determination of the conformation of cyclic and linear model peptides - Studies on the chiral dirhodium complexes by ECD and VCD Protein NMR and protein modeling - Computational work on peptides. - NMR studies in proteins Protein X-ray crystallography - The crystal structures of prolyl oligopeptidase (POP) in complexes were solved. - Studies on calmodulin (CaM). - Structural aspects of molecular recognition were characterized. - The crystal structure of Pyrococcus horikoshii acylaminoacyl peptidase was solved. Biochemical studies - The structure of a scallop peptide in three crystal environments was determined . - The structure of the tail domain of myosin-6 was predicted. - A thorough analysis of cross-predictions of coiled-coil and disordered protein segments was performed. - Microsecond classical molecular dynamics simulations of five naturally occurring and two synthetic charged single ?-helices were performed. - Studies on LC8 dynein light chain (DYNLL), a hub protein. Immunological studies - Studies on SHP-2 tyrosine phosphatase. A variety of phosphopeptides were tested. - Structural studies were performed on biotinylated collagen epitope peptide, an extravidin-peptide complex and extravidin. Papers: 3

Elsődleges genetikai vizsgálat Prader–Willi-szindróma igazolására

INTRODUCTION: According to the international literature, DNA methylation analysis of the promoter region of SNRPN locus is the most efficient way to start genetic investigation in patients with suspected Prader-Willi syndrome. AIM: Our aim was to develop a simple, reliable first-tier diagnosis to confirm Prader-Willi syndrome, therefore to compare our self-designed simple, cost-efficient high-resolution melting analysis and the most commonly used methylation-specific multiplex ligation-dependent probe amplification to confirm Prader-Willi syndrome. METHOD: We studied 17 clinically suspected Prader-Willi syndrome children and their DNA samples. With self-designed primers, bisulfite-sensitive polymerase chain reaction, high-resolution melting analysis and, as a control, methylation-specific multiplex ligation-dependent probe amplification were performed. RESULTS: Prader-Willi syndrome was genetically confirmed in 6 out of 17 clinically suspected Prader-Willi syndrome patients. The results of high-resolution melting analysis and methylation-specific multiplex ligation-dependent probe amplification were equivalent in each case. CONCLUSION: Using our self-designed primers and altered bisulfite-specific PCR conditions, high-resolution melting analysis appears to be a simple, fast, reliable and effective method for primarily proving or excluding clinically suspected Prade-Willi syndrome cases. Orv Hetil. 2018; 159(2): 64-69

Comprehensive DNA Methylation Analysis Reveals a Common Ten-Gene Methylation Signature in Colorectal Adenomas and Carcinomas

Microarray analysis of promoter hypermethylation provides insight into the role and extent of DNA methylation in the development of colorectal cancer (CRC) and may be co-monitored with the appearance of driver mutations. Colonic biopsy samples were obtained endoscopically from 10 normal, 23 adenoma (17 low-grade (LGD) and 6 high-grade dysplasia (HGD)), and 8 ulcerative colitis (UC) patients (4 active and 4 inactive). CRC samples were obtained from 24 patients (17 primary, 7 metastatic (MCRC)), 7 of them with synchronous LGD. Field effects were analyzed in tissues 1 cm (n = 5) and 10 cm (n = 5) from the margin of CRC. Tissue materials were studied for DNA methylation status using a 96 gene panel and for KRAS and BRAF mutations. Expression levels were assayed using whole genomic mRNA arrays. SFRP1 was further examined by immunohistochemistry. HT29 cells were treated with 5-aza-2' deoxycytidine to analyze the reversal possibility of DNA methylation. More than 85% of tumor samples showed hypermethylation in 10 genes (SFRP1, SST, BNC1, MAL, SLIT2, SFRP2, SLIT3, ALDH1A3, TMEFF2, WIF1), whereas the frequency of examined mutations were below 25%. These genes distinguished precancerous and cancerous lesions from inflamed and healthy tissue. The mRNA alterations that might be caused by systematic methylation could be partly reversed by demethylation treatment. Systematic changes in methylation patterns were observed early in CRC carcinogenesis, occuring in precursor lesions and CRC. Thus we conclude that DNA hypermethylation is an early and systematic event in colorectal carcinogenesis, and it could be potentially reversed by systematic demethylation therapy, but it would need more in vitro and in vivo experiments to support this theory

Syndecan-1 and FGF-2, but Not FGF Receptor-1, Share a Common Transport Route and Co-Localize with Heparanase in the Nuclei of Mesenchymal Tumor Cells

Syndecan-1 forms complexes with growth factors and their cognate receptors in the cell membrane. We have previously reported a tubulin-mediated translocation of syndecan-1 to the nucleus. The transport route and functional significance of nuclear syndecan-1 is still incompletely understood. Here we investigate the sub-cellular distribution of syndecan-1, FGF-2, FGFR-1 and heparanase in malignant mesenchymal tumor cells, and explore the possibility of their coordinated translocation to the nucleus. To elucidate a structural requirement for this nuclear transport, we have transfected cells with a syndecan-1/EGFP construct or with a short truncated version containing only the tubulin binding RMKKK sequence. The sub-cellular distribution of the EGFP fusion proteins was monitored by fluorescence microscopy. Our data indicate that syndecan-1, FGF-2 and heparanase co-localize in the nucleus, whereas FGFR-1 is enriched mainly in the perinuclear area. Overexpression of syndecan-1 results in increased nuclear accumulation of FGF-2, demonstrating the functional importance of syndecan-1 for this nuclear transport. Interestingly, exogenously added FGF-2 does not follow the route taken by endogenous FGF-2. Furthermore, we prove that the RMKKK sequence of syndecan-1 is necessary and sufficient for nuclear translocation, acting as a nuclear localization signal, and the Arginine residue is vital for this localization. We conclude that syndecan-1 and FGF-2, but not FGFR-1 share a common transport route and co-localize with heparanase in the nucleus, and this transport is mediated by the RMKKK motif in syndecan-1. Our study opens a new perspective in the proteoglycan field and provides more evidence of nuclear interactions of syndecan-1

Remodeling of extracellular matrix by normal and tumor-associated fibroblasts promotes cervical cancer progression

Background: Comparison of tissue microarray results of 29 cervical cancer and 27 normal cervix tissue samples using immunohistochemistry revealed considerable reorganization of the fibrillar stroma of these tumors. Preliminary densitometry analysis of laminin-1, α -smooth muscle actin (SMA) and fibronectin immunostaining demonstrated 3.8-fold upregulation of laminin-1 and 5.2-fold increase of SMA in the interstitial stroma, indicating that these proteins and the activated fibroblasts play important role in the pathogenesis of cervical cancer. In the present work we investigated the role of normal and tumor-associated fibroblasts. Methods: In vitro models were used to throw light on the multifactorial process of tumor-stroma interaction, by means of studying the cooperation between tumor cells and fibroblasts. Fibroblasts from normal cervix and cervical cancers were grown either separately or in co-culture with CSCC7 cervical cancer cell line. Changes manifest in secreted glycoproteins, integrins and matrix metallo-proteases (MMPs) were explored. Results: While normal fibroblasts produced components of interstitial matrix and TGF- β 1 that promoted cell proliferation, cancer-associated fibroblasts (CAFs) synthesized ample amounts of laminin-1. The following results support the significance of laminin-1 in the invasion of CSCC7 cells: 1.) Tumor-associated fibroblasts produced more laminin-1 and less components of fibrillar ECM than normal cells; 2.) The production of laminin chains was further increased when CSCC7 cells were grown in co-culture with fibroblasts; 3.) CSCC7 cells were capable of increasing their laminin production; 4.) Tumor cells predominantly expressed integrin α 6 β 4 laminin receptors and migrated towards laminin. The integrin profile of both normal and tumor-associated fibroblasts was similar, expressing receptors for fibronectin, vitronectin and osteopontin. MMP-7 secreted by CSCC7 cells was upregulated by the presence of normal fibroblasts, whereas MMP-2 produced mainly by fibroblasts was activated in the presence of CSCC7 cells. Conclusions: Our results indicate that in addition to degradation of the basement membrane, invasion of cervical cancer is accomplished by the remodeling of the interstitial stroma, which process includes decrease and partial replacement of fibronectin and collagens by a laminin-rich matrix

Syndecan-1 Enhances Proliferation, Migration and Metastasis of HT-1080 Cells in Cooperation with Syndecan-2

Syndecans are transmembrane heparan sulphate proteoglycans. Their role in the development of the malignant phenotype is ambiguous and depends upon the particular type of cancer. Nevertheless, syndecans are promising targets in cancer therapy, and it is important to elucidate the mechanisms controlling their various cellular effects. According to earlier studies, both syndecan-1 and syndecan-2 promote malignancy of HT-1080 human fibrosarcoma cells, by increasing the proliferation rate and the metastatic potential and migratory ability, respectively. To better understand their tumour promoter role in this cell line, syndecan expression levels were modulated in HT-1080 cells and the growth rate, chemotaxis and invasion capacity were studied. For in vivo testing, syndecan-1 overexpressing cells were also inoculated into mice. Overexpression of full length or truncated syndecan-1 lacking the entire ectodomain but containing the four juxtamembrane amino acids promoted proliferation and chemotaxis. These effects were accompanied by a marked increase in syndecan-2 protein expression. The pro-migratory and pro-proliferative effects of truncated syndecan-1 were not observable when syndecan-2 was silenced. Antisense silencing of syndecan-2, but not that of syndecan-1, inhibited cell migration. In vivo, both full length and truncated syndecan-1 increased tumour growth and metastatic rate. Based on our in vitro results, we conclude that the tumour promoter role of syndecan-1 observed in HT-1080 cells is independent of its ectodomain; however, in vivo the presence of the ectodomain further increases tumour proliferation. The enhanced migratory ability induced by syndecan-1 overexpression is mediated by syndecan-2. Overexpression of syndecan-1 also leads to activation of IGF1R and increased expression of Ets-1. These changes were not evident when syndecan-2 was overexpressed. These findings suggest the involvement of IGF1R and Ets-1 in the induction of syndecan-2 synthesis and stimulation of proliferation by syndecan-1. This is the first report demonstrating that syndecan-1 enhances malignancy of a mesenchymal tumour cell line, via induction of syndecan-2 expression

A versenyképes mezőgazdaság, a földár és a föld jövedelemtermelő képességének összefüggései

Az EU-csatlakozás óta jelentős mértékben nőtt a kifizetett támogatások összege, ezzel együtt pedig az ágazat jövedelme. A jövőben a támogatások helyett a versenyképesség kerül előtérbe, ez pedig komolyabb feladat, mint a támogatások kiharcolása és fenntartása. A támogatások fokozatos leépítése elsősorban a közvetlen területalapú támogatásoknál várható. Ugyanakkor erős korreláció tapasztalható a földbérleti díjak emelkedése és a növekvő közvetlen támogatások között. Ezzel szemben a beruházások bővülése elmaradt a jövedelem növekedésének ütemétől, mert azokat leginkább a célzott fejlesztési támogatások befolyásolták. A közvetlen kifizetések szinte egésze a magasabb földárban tőkésül, aminek következtében nő a földvásárlás vagy földbérlet költsége. A földtulajdonos is részesedik a támogatásból, ezáltal emelkedik a földtulajdonnal rendelkező gazdák vagyona is, de a következő gazdálkodói generáció nagyobb tőkét és/vagy működési költséget és alacsonyabb termelési hatékonyságot örököl. Megállapítható, hogy a föld értékét sokkal inkább meghatározza a közgazdasági környezet, mint maga a föld piaci jövedelemtermelő képessége. Az árutőzsdei árak egyelőre nem igazolják vissza a precíziós gazdálkodás többletértékét. A precíziós gazdálkodásnál alacsonyabb üzemanyagköltséggel, ezzel párhuzamosan csökkenő környezetterheléssel lehet számolni, de ezt ma nem feltétlenül fizeti meg a vevő, nem ismeri el többletértékként az árutőzsde. A precíziós növénytermelési technológia eredményes működtetéséhez nem elégséges csupán a beruházás pénzügyi fedezetének biztosítása, szükség van a gazdálkodó, illetőleg a munkafolyamatokban részt vevő munkaerő aktív részvételére és pozitív hozzáállására. A sikeres gazdálkodás feltétele ma a képzés, a fejlesztés, az innováció, az alkalmazkodóképesség és a munkavállalók motiválása. = Since the accession of Hungary to the European Union, the amount of subsidies paid to agriculture has increased considerably, as has the income of the sector. However, the gradual phasing out of direct area payments can be expected. At the same time, there is a strong correlation between rising land lease fees and increasing direct subsidies. By contrast, investment growth lagged behind the rate of increase in agricultural income because investments were most affected by targeted rural development subsidies. In the case of direct payments, subsidies are almost entirely capitalised in higher land prices, resulting in rising cost of land acquisition or land lease. Landowners also benefit from the subsidies, thus increasing the wealth of land owners, but the next generation inherits greater capital and / or operating costs, thus lower production efficiency. The value of land is determined more by the economic environment than by the land itself. Commodity prices do not yet justify the added value of precision farming. The lower input use (fuel, fertiliser, chemicals etc.) and decreasing environmental impact of precision farming are not necessarily recognised by higher commodity prices as added value. In addition to the financial coverage of investment, the active participation and positive attitude of the farmer is necessary to ensure the successful operation of precision farming. The conditions of successful farm management are training, development, innovation, adaptability and motivation of employees