Promoter Methylation of LEP and LEPR before and after Bariatric Surgery: A Cross-Sectional Study

IntroductionDNA methylation constitutes one important epigenetic mechanism that regulates gene expression in human cells. With regard to obesity, bariatric surgery-induced weight loss has been associated with promoter methylation changes in several genes. Hyperleptinemia is a characteristic feature of obesity. The underlying regulating mechanisms have not yet been completely elucidated.MethodsWe investigated the methylation of the promoters of the leptin gene (LEP) and the leptin receptor gene (LEPR) as well as leptin expression in pre- and postbariatric surgery patients using a comparative cross-sectional design.ResultsOur results revealed significantly higher LEP promoter methylation patterns in prebariatric surgery patients compared to postoperatively. DNA methylation of the LEPR promoter was significantly higher in the postoperative group. Moreover, we found significantly higher leptin serum levels in patients before the bariatric surgery than afterwards.DiscussionThese findings strengthen the suggestion that there is an association between LEP expression and LEP methylation in obesity. We suggest that the epigenetic profile of LEP might be influenced by leptin serum levels in the form of a regulating feedback mechanism.     

Roles of mechano-sensitive ion channels, cytoskeleton, and contractile activity in stretch-induced immediate-early gene expression and hypertrophy of cardiac myocytes.

Mechanical loading of cardiac and skeletal muscles in vivo and in vitro causes rapid activation of a number of immediate-early (IE) genes and hypertrophy of muscle cells. However, little is known as to how muscle cells sense mechanical load and transduce it into intracellular signals of gene regulation. We examined roles of putative cellular mechanotransducers, mechanosensitive ion channels, the cytoskeleton, and contractile activity in stretch-induced hypertrophy of cardiac myocytes grown on a deformable silicone sheet. Using the patch-clamp technique, we found a single class of stretch-activated cation channel that was completely blocked by gadolinium (Gd3+). Inhibition of this channel by Gd3+ did not affect either the stretch-induced expression of IE genes or the increase in protein synthesis. Neither disruption of microtubules with colchicine nor that of actin microfilaments by cytochalasin D prevented the stretch-induced IE gene expression and increase in protein synthesis. Arresting contractile activity of myocytes by high K+, tetrodotoxin, or Ba2+ did not affect the stretch-induced IE gene expression. Tetrodotoxin-arrested myocytes could increase protein synthesis in response to stretch. These results suggest that Gd(3+)-sensitive ion channels, microtubules, microfilaments, and contractile activity may not be necessary for transduction of mechanical stretch into the IE gene expression and hypertrophy. The stimulus of membrane stretch may be transmitted to the cell nucleus through some mechanisms other than electrical or direct mechanical transduction in cardiac myocytes.     

Accuracy of supplementary serologic testing for human T-lymphotropic virus types I and II in US blood donors. Retrovirus Epidemiology Donor Study

Blood donations in the United States have been screened for antibody to human T-lymphotropic virus type I (HTLV-I) by HTLV-I enzyme immunoassay (EIA) since November 1988. Specimens repeatedly found to be reactive by EIA undergo confirmation by supplementary serologic tests. We assessed the accuracy of blood center testing of 994 HTLV-I EIA repeat-reactive specimens in five US blood centers between November 1988 and December 1991. Of 410 confirmed HTLV-I/II donations, 407 (99.3%) were infected with HTLV-I/II, as determined by polymerase chain reaction (PCR) (403 cases) and by repeat serologic testing (4 cases). The three false- positive results occurred in the first year of testing. Of 425 HTLV- indeterminate specimens, 6 (1.4%) were found to be infected by PCR (5 with HTLV-II and 1 with HTLV-I). None of 159 confirmatory test-negative donations was PCR positive. Of HTLV-I/II-seropositive specimens, 80.2% to 95.4% could be typed as HTLV-I or HTLV-II by type-specific serologic assays. These results support recommendations that HTLV-I/II- seropositive donors should be advised that they are infected with HTLV- I, HTLV-II, or HTLV-I/II (depending on results of type-specific assays). HTLV-indeterminate donors should be advised that their results only rarely indicate HTLV infection. HTLV confirmatory test-negative donors should be reassured that they are not infected with HTLV-I or HTLV-II.     

Copy number variants in patients with short stature

Height is a highly heritable and classic polygenic trait. Recent genome-wide association studies (GWAS) have revealed that at least 180 genetic variants influence adult height. However, these variants explain only about 10% of the phenotypic variation in height. Genetic analysis of short individuals can lead to the discovery of novel rare gene defects with a large effect on growth. In an effort to identify novel genes associated with short stature, genome-wide analysis for copy number variants (CNVs), using single-nucleotide polymorphism arrays, in 162 patients (149 families) with short stature was performed. Segregation analysis was performed if possible, and genes in CNVs were compared with information from GWAS, gene expression in rodents'' growth plates and published information. CNVs were detected in 40 families. In six families, a known cause of short stature was found (SHOX deletion or duplication, IGF1R deletion), in two combined with a de novo potentially pathogenic CNV. Thirty-three families had one or more potentially pathogenic CNVs (n=40). In 24 of these families, segregation analysis could be performed, identifying three de novo CNVs and nine CNVs segregating with short stature. Four were located near loci associated with height in GWAS (ADAMTS17, TULP4, PRKG2/BMP3 and PAPPA). Besides six CNVs known to be causative for short stature, 40 CNVs with possible pathogenicity were identified. Segregation studies and bioinformatics analysis suggested various potential candidate genes.     

Phosphorylation-regulated axonal dependent transport of syntaxin 1 is mediated by a Kinesin-1 adapter

Presynaptic nerve terminals are formed from preassembled vesicles that are delivered to the prospective synapse by kinesin-mediated axonal transport. However, precisely how the various cargoes are linked to the motor proteins remains unclear. Here, we report a transport complex linking syntaxin 1a (Stx) and Munc18, two proteins functioning in synaptic vesicle exocytosis at the presynaptic plasma membrane, to the motor protein Kinesin-1 via the kinesin adaptor FEZ1. Mutation of the FEZ1 ortholog UNC-76 in Caenorhabditis elegans causes defects in the axonal transport of Stx. We also show that binding of FEZ1 to Kinesin-1 and Munc18 is regulated by phosphorylation, with a conserved site (serine 58) being essential for binding. When expressed in C. elegans, wild-type but not phosphorylation-deficient FEZ1 (S58A) restored axonal transport of Stx. We conclude that FEZ1 operates as a kinesin adaptor for the transport of Stx, with cargo loading and unloading being regulated by protein kinases.     

Temporomandibular Joint: MR Fast Scanning

Eleven hundred ninety temporomandibular joints (TMJ) in 662 patients were studied with high field strength surface coil magnetic resonance using high resolution T1-weighted closed-mouth and short partial flip angle closed- and open-mouth images to evaluate internal derangement. Partial flip angle (gradient recalled acquisition in the steady state, or GRASS) images using image acquisition times as short as three seconds in most cases provided reliable information on disk position, morphology, and intrinsic signal. Abnormalities of disk function may be studied using sequential GRASS scans during incremental degrees of mouth opening. Joint effusions may be delineated by T2* effects, and avascular necrosis (AVN) of the mandibular condyle is demonstrated. Fast GRASS scans provide accurate information and may be employed alone or in conjunction with standard T1-weighted images in studies of the TMJ. Combined studies using both T1-weighted closed- and short GRASS open-mouth scans provide greater information than is obtained from either T1-weighted or GRASS scans alone, and examination times are short compared to T2-weighted pulse sequence techniques.     

CD44 enhances tumor formation and lung metastasis in experimental osteosarcoma and is an additional predictor for poor patient outcome

Formation of metastases in the lungs is the major cause of death in patients suffering from osteosarcoma (OS). Metastases at presentation and poor response to preoperative chemotherapy are strong predictors for poor patient outcome. The elucidation of molecular markers that promote metastasis formation and/or chemoresistance is therefore of importance. CD44 is a plasma membrane glycoprotein that binds to the extracellular matrix component hyaluronan (HA) and has been shown to be involved in metastasis formation in a variety of other tumors. Here we investigated the role of CD44 expression on OS tumor formation and metastasis. High CD44 expression, evaluated with a tissue microarray including samples from 53 OS patients and stained with a pan‐CD44 antibody (Hermes3), showed a tendency (p < 0.08) to shortened overall survival. However, nonresponders and patients with lung metastases and high CD44 expression had significantly poorer prognosis than patients with low CD44 expression. Overexpression of the standard CD44 isoform (CD44s) and its HA‐binding defective mutant R41A in osteoblastic SaOS‐2 cells resulted in HA‐independent higher migration rates and increased chemoresistance, partially dependent on HA. In an orthotopic mouse model of OS, overexpression of CD44s in SaOS‐2 cells resulted in an HA‐dependent increased primary tumor formation and increased numbers of micrometastases and macrometastases in the lungs. In conclusion, although CD44 failed to be an independent predictor for patient outcome in this limited cohort of OS patients, increased CD44 expression was associated with even worse survival in patients with chemoresistance and with lung metastases. CD44‐associated chemoresistance was also observed in vitro, and increased formation of lung metastases was found in vivo in SCID mice. © 2013 American Society for Bone and Mineral Research.