Helicases are motor proteins that couple the hydrolysis of nucleoside triphosphate (NTPase) to nucleic acid unwinding. The hexameric helicases have a characteristic ring-shaped structure, and all, except the eukaryotic minichromosomal maintenance (MCM) helicase, are homohexamers. Most of the 12 known hexameric helicases play a role in DNA replication, recombination, and transcription. A human genetic disorder, Bloom's syndrome, is associated with a defect in one member of the class of hexameric helicases. Significant progress has been made in understanding the biochemical properties, structures, and interactions of these helicases with DNA and nucleotides. Cooperativity in nucleotide binding was observed in many, and sequential NTPase catalysis has been observed in two proteins, gp4 of bacteriophage T7 and rho of Escherichia coli. The crystal structures of the oligomeric T7 gp4 helicase and the hexamer of RepA helicase show structural features that substantiate the observed cooperativity, and both are consistent with nucleotide binding at the subunit interface. Models are presented that show how sequential NTP hydrolysis can lead to unidirectional and processive translocation. Possible unwinding mechanisms based on the DNA exclusion model are proposed here, termed the wedge, torsional, and helix-destabilizing models.

Objective To study the longitudinal rate of (and sensitivity to) change of knee cartilage thickness across defined stages of radiographic osteoarthritis (ROA), specifically healthy knees and knees with end-stage ROA. Methods One knee of 831 Osteoarthritis Initiative (OAI) participants was examined: 112 healthy, without ROA or risk factors for knee OA, and 719 ROA knees: 310 calculated Kellgren Lawrence [cKLG] grade 2, 300 cKLG3, and 109 cKLG4. Subregional change in thickness was assessed after segmentation of weight-bearing femorotibial cartilage at baseline and at one year from coronal MRI. Regional and ordered values (OV) of change were compared by baseline ROA status. Results Healthy knees displayed small changes in plates and subregions (±0.7%; standardized response mean [SRM] ±0.15), with OVs being symmetrically distributed around zero. In cKLG2 knees, changes in cartilage thickness were small (≤1%; minimal SRM -0.22) and not significantly different from healthy knees. Knees with cKLG3 showed substantial loss of cartilage thickness (up to -2.5%; minimal SRM -0.35), with OV changes being significantly (p<0.05) greater than those in healthy knees. cKLG4 knees displayed the largest rate of loss across ROA grades (up to -3.9%; minimal SRM -0.51), with OV changes also significantly (p<0.05) greater than in healthy knees. Conclusion MRI-based cartilage thickness showed high rates of loss in knees with moderate and end-stage ROA, and small rates (indistinguishable from healthy knees) in mild ROA. From the perspective of sensitivity to change, end-stage ROA knees need not be excluded from longitudinal studies using MRI cartilage morphology as an endpoint.

Objective The Osteoarthritis Initiative (OAI) is targeted at identifying sensitive biomarkers and risk factors of symptomatic knee OA onset and progression. Quantitative cartilage imaging in the OAI relies on validated FLASH sequences that suffer from relatively long acquisition times, and on a near-isotropic double echo steady state (DESS) sequence. We therefore directly compared the sensitivity to cartilage thickness changes and the correlation of these protocols longitudinally. Methods Baseline and 12 month followup data of 80 knees were acquired using 1.5mm coronal FLASH and 0.7mm sagittal DESS sequences. In these and in 1.5mm coronal multiplanar reconstructions (MPR) of the DESS the medial femorotibial cartilage was segmented with blinding to acquisition order. In the weight-bearing femoral condyle, a 60% (distance trochlear notch to posterior femur) and 75% region of interest (ROI) were studied. Results The standardized response mean (SRM=mean change /standard deviation of change) in central medial femorotibial (cMFTC) cartilage thickness was −0.34 for coronal FLASH, −0.37 for coronal MPR DESS, −0.36 for sagittal DESS with the 60% ROI, and −0.38 for the 75% ROI. Using every 2nd 0.7mm sagittal slice (DESS) yielded similar SRMs in cMFTC for the 60% and 75% ROI from odd (−0.35/−0.36) and even slice numbers (−0.36/−0.39), respectively. Baseline cartilage thickness displayed high correlations (r≥0.94) between the three protocols; the correlations of longitudinal changes were ≥0.79 (Pearson) and ≥0.45 (Spearman). Conclusions Cartilage morphometry with FLASH and DESS display similar longitudinal sensitivity to change. Analysis of every second slice of the 0.7mm DESS provides adequate sensitivity to change.

ObjectiveIn addition to improve glucose intolerance, recent studies suggest that glucagon-like peptide-1 (GLP-1) receptor agonism also decreases triglyceride (TG) levels. The aim of this study was to evaluate the effect of GLP-1 receptor agonism on very-low-density lipoprotein (VLDL)-TG production and liver TG metabolism.Experimental ApproachThe GLP-1 peptide analogues CNTO3649 and exendin-4 were continuously administered subcutaneously to high fat diet-fed APOE*3-Leiden transgenic mice. After 4 weeks, hepatic VLDL production, lipid content, and expression profiles of selected genes involved in lipid metabolism were determined.ResultsCNTO3649 and exendin-4 reduced fasting plasma glucose (up to −30% and −28% respectively) and insulin (−43% and −65% respectively). In addition, these agents reduced VLDL-TG production (−36% and −54% respectively) and VLDL-apoB production (−36% and −43% respectively), indicating reduced production of VLDL particles rather than reduced lipidation of apoB. Moreover, they markedly decreased hepatic content of TG (−39% and −55% respectively), cholesterol (−30% and −55% respectively), and phospholipids (−23% and −36% respectively), accompanied by down-regulation of expression of genes involved in hepatic lipogenesis (Srebp-1c, Fasn, Dgat1) and apoB synthesis (Apob).ConclusionGLP-1 receptor agonism reduces VLDL production and hepatic steatosis in addition to an improvement of glycemic control. These data suggest that GLP-receptor agonists could reduce hepatic steatosis and ameliorate dyslipidemia in patients with type 2 diabetes mellitus.

These data suggest that pretreatment of islets and the recipient with anti-TF may limit the effects of IBMIR, thereby enhancing islet engraftment and survival.