Tobramycin exposure from active calcium sulfate bone graft substitute.

BACKGROUND: Bone graft substitute such as calcium sulfate are frequently used as carrier material for local antimicrobial therapy in orthopedic surgery. This study aimed to assess the systemic absorption and disposition of tobramycin in patients treated with a tobramycin-laden bone graft substitute (Osteoset® T). METHODS: Nine blood samples were taken from 12 patients over 10 days after Osteoset® T surgical implantation. Tobramycin concentration was measured by fluorescence polarization. Population pharmacokinetic analysis was performed using NONMEM to assess the average value and variability (CV) of pharmacokinetic parameters. Bioavailability (F) was assessed by equating clearance (CL) with creatinine clearance (Cockcroft CLCr). Based on the final model, simulations with various doses and renal function levels were performed. (ClinicalTrials.gov number, NCT01938417). RESULTS: The patients were 52 +/- 20 years old, their mean body weight was 73 +/- 17 kg and their mean CLCr was 119 +/- 55 mL/min. Either 10 g or 20 g Osteoset® T with 4% tobramycin sulfate was implanted in various sites. Concentration profiles remained low and consistent with absorption rate-limited first-order release, while showing important variability. With CL equated to CLCr, mean absorption rate constant (ka) was 0.06 h-1, F was 63% or 32% (CV 74%) for 10 and 20 g Osteoset® T respectively, and volume of distribution (V) was 16.6 L (CV 89%). Simulations predicted sustained high, potentially toxic concentrations with 10 g, 30 g and 50 g Osteoset® T for CLCr values below 10, 20 and 30 mL/min, respectively. CONCLUSIONS: Osteoset® T does not raise toxicity concerns in subjects without significant renal failure. The risk/benefit ratio might turn unfavorable in case of severe renal failure, even after standard dose implantation.

Neurogenic subventricular zone stem/progenitor cells are notch1-dependent in their active but not quiescent state.

The adult mammalian forebrain contains neural stem/progenitor cells (NSCs) that generate neurons throughout life. As in other somatic stem cell systems, NSCs are proposed to be predominantly quiescent and proliferate only sporadically to produce more committed progeny. However, quiescence has recently been shown not to be an essential criterion for stem cells. It is not known whether NSCs show differences in molecular dependence based on their proliferation state. The subventricular zone (SVZ) of the adult mouse brain has a remarkable capacity for repair by activation of NSCs. The molecular interplay controlling adult NSCs during neurogenesis or regeneration is not clear but resolving these interactions is critical in order to understand brain homeostasis and repair. Using conditional genetics and fate mapping, we show that Notch signaling is essential for neurogenesis in the SVZ. By mosaic analysis, we uncovered a surprising difference in Notch dependence between active neurogenic and regenerative NSCs. While both active and regenerative NSCs depend upon canonical Notch signaling, Notch1-deletion results in a selective loss of active NSCs (aNSCs). In sharp contrast, quiescent NSCs (qNSCs) remain after Notch1 ablation until induced during regeneration or aging, whereupon they become Notch1-dependent and fail to fully reinstate neurogenesis. Our results suggest that Notch1 is a key component of the adult SVZ niche, promoting maintenance of aNSCs, and that this function is compensated in qNSCs. Therefore, we confirm the importance of Notch signaling for maintaining NSCs and neurogenesis in the adult SVZ and reveal that NSCs display a selective reliance on Notch1 that may be dictated by mitotic state.

Estimating dormant and active hematopoietic stem cell kinetics through extensive modeling of bromodeoxyuridine label-retaining cell dynamics.

Bone marrow hematopoietic stem cells (HSCs) are responsible for both lifelong daily maintenance of all blood cells and for repair after cell loss. Until recently the cellular mechanisms by which HSCs accomplish these two very different tasks remained an open question. Biological evidence has now been found for the existence of two related mouse HSC populations. First, a dormant HSC (d-HSC) population which harbors the highest self-renewal potential of all blood cells but is only induced into active self-renewal in response to hematopoietic stress. And second, an active HSC (a-HSC) subset that by and large produces the progenitors and mature cells required for maintenance of day-to-day hematopoiesis. Here we present computational analyses further supporting the d-HSC concept through extensive modeling of experimental DNA label-retaining cell (LRC) data. Our conclusion that the presence of a slowly dividing subpopulation of HSCs is the most likely explanation (amongst the various possible causes including stochastic cellular variation) of the observed long term Bromodeoxyuridine (BrdU) retention, is confirmed by the deterministic and stochastic models presented here. Moreover, modeling both HSC BrdU uptake and dilution in three stages and careful treatment of the BrdU detection sensitivity permitted improved estimates of HSC turnover rates. This analysis predicts that d-HSCs cycle about once every 149-193 days and a-HSCs about once every 28-36 days. We further predict that, using LRC assays, a 75%-92.5% purification of d-HSCs can be achieved after 59-130 days of chase. Interestingly, the d-HSC proportion is now estimated to be around 30-45% of total HSCs - more than twice that of our previous estimate.

Report on a review of selected application controls over the University of Northern Iowa General Ledger System for the period of June 22, 2011 through August 4, 2011

Report on a review of selected application controls over the University of Northern Iowa General Ledger System for the period of June 22, 2011 through August 4, 2011

Intracellular ubiquitylation of the epithelial Na+ channel controls extracellular proteolytic channel activation via conformational change.

The epithelial Na(+) channel ENaC is a key player in the maintenance of whole body Na(+) balance, and consequently of blood pressure. It is tightly regulated by numerous signaling pathways including ubiquitylation via the ubiquitin-protein ligase Nedd4-2. This mechanism is itself under the control of several kinases, which phosphorylate Nedd4-2, thereby interfering with ENaC/Nedd4-2 interaction, or by Usp2-45, which binds to and deubiquitylates ENaC. Another, different regulatory mechanism concerns the proteolytic activation of ENaC, during which the channel is cleaved on its luminal side by intracellular convertases such as furin, and further activated by extracellular proteases such as CAP-1. This process is regulated as well but the underlying mechanisms are not understood. Previously, evidence was provided that the ubiquitylation status of ENaC may affect the cleavage of the channel. When ubiquitylation of ENaC was reduced, either by co-expressing Usp2-45, or mutating either the ENaC PY-motifs (i.e. the binding sites for Nedd4-2) or intracellular lysines (i.e. ubiquitylation sites), the level of channel cleavage was increased. Here we demonstrate that lysine-mutated ENaC channels are not ubiquitylated at the cell surface, are preferentially cleaved, and Usp2-45 does not affect their cleavage efficiency. We further show by limited proteolysis that the intracellular ubiquitylation status of ENaC affects the extracellular conformation of αENaC, by demonstrating that non-ubiquitylated channels are more efficiently cleaved when treated with extracellularly added trypsin or chymotrypsin. These results present a new paradigm in which an intracellular, post-translational modification (e.g. ubiquitylation) of a transmembrane protein can affect its extracellular conformation.

Active uptake of dendritic cell-derived exovesicles by epithelial cells induces the release of inflammatory mediators through a TNF-alpha-mediated pathway.

Dendritic cells (DCs) can release hundreds of membrane vesicles, called exovesicles, which are able to activate resting DCs and distribute antigen. Here, we examined the role of mature DC-derived exovesicles in innate and adaptive immunity, in particular their capacity to activate epithelial cells. Our analysis of exovesicle contents showed that exovesicles contain major histocompatibility complex-II, CD40, and CD83 molecules in addition to tumor necrosis factor (TNF) receptors, TNFRI and TNFRII, and are important carriers of TNF-alpha. These exovesicles are rapidly internalized by epithelial cells, inducing the release of cytokines and chemokines, but do not transfer an alloantigen-presenting capacity to epithelial cells. Part of this activation appears to involve the TNF-alpha-mediated pathway, highlighting the key role of DC-derived exovesicles, not only in adaptive immunity, but also in innate immunity by triggering innate immune responses and activating neighboring epithelial cells to release cytokines and chemokines, thereby amplifying the magnitude of the innate immune response.

Multi-Atlas based Segmentation of Head and Neck CT Images using Active Contour

This paper presents the segmentation of bilateral parotid glands in the Head and Neck (H&N) CT images using an active contour based atlas registration. We compare segmentation results from three atlas selection strategies: (i) selection of "single-most-similar" atlas for each image to be segmented, (ii) fusion of segmentation results from multiple atlases using STAPLE, and (iii) fusion of segmentation results using majority voting. Among these three approaches, fusion using majority voting provided the best results. Finally, we present a detailed evaluation on a dataset of eight images (provided as a part of H&N auto segmentation challenge conducted in conjunction with MICCAI-2010 conference) using majority voting strategy.

Constitutively active mutants of the beta1-adrenergic receptor.

We provide the first evidence that point mutations can constitutively activate the beta(1)-adrenergic receptor (AR). Leucine 322 of the beta(1)-AR in the C-terminal portion of its third intracellular loop was replaced with seven amino acids (I, T, E, F, C, A and K) differing in their physico-chemical properties. The beta(1)-AR mutants expressed in HEK-293 cells displayed various levels of constitutive activity which could be partially inhibited by some beta-blockers. The results of this study might have interesting implications for future studies aiming at elucidating the activation process of the beta(1)-AR as well as the mechanism of action of beta-blockers.

Report on a review of selected application controls over the Iowa Department of Transportation’s Driver’s License System for the period May 2, 2011 through May 27, 2011

Report on a review of selected application controls over the Iowa Department of Transportation’s Driver’s License System for the period May 2, 2011 through May 27, 2011

Report on a review of selected application controls over the Iowa State University of Science and Technology Facilities Planning and Management - Facilities Administrative Management Information System for the period of April 18, 2011 through May 16, 2011

Report on a review of selected application controls over the Iowa State University of Science and Technology Facilities Planning and Management - Facilities Administrative Management Information System for the period of April 18, 2011 through May 16, 2011

Report on a review of selected general and application controls over the Iowa Department of Human Services’ Medicaid Management Information System for the period April 4, 2011 through April 29, 2011

Report on a review of selected general and application controls over the Iowa Department of Human Services’ Medicaid Management Information System for the period April 4, 2011 through April 29, 2011

Body composition using bio-impedance analysis in pediatric patients with inflammatory bowel disease. Corcordance with dual energy X-ray absorptiometry and comparison with healthy controls

Introduction: Growth is a central process in paediatrics. Weight and height evaluation are therefore routine exams for every child but in some situation, particularly inflammatory bowel disease (IBD), a wider evaluation of nutritional status needs to be performed. Objectives: To assess the accuracy of bio-impedance analysis (BIA) compared to the gold standard dual energy X-ray absorptiometry (DEXA) in estimating percentage body fat (fat mass; FM) and lean body mass (fat free mass; FFM) in children with inflammatory bowel disease (IBD). To compare FM and FFM levels between patients with IBD and healthy controls. Methods: Twenty-nine healthy controls (12 females; mean age: 12.7 ± 1.9 years) and 21 patients (11 females; 14.3 ± 1.3 years) were recruited from August 2011 to October 2012 at our institution. BIA was performed in all children and DEXA in patients only. Concordance between BIA and DEXA was assessed using Lin's concordance correlation and the Bland-Altman method. Between-group comparisons were made using analysis of variance adjusting for age. Results: BIA-derived FM% showed a good concordance with DEXA-derived values, while BIA-derived FFM% tended to be slightly higher than DEXA-derived values (table). No differences were found between patients and controls regarding body mass index (mean ± SD: 19.3 ± 3.3 vs. 20.1 ± 2.8 kg/m2, respectively; age-adjusted P = 0.08) and FM% (boys: 25.3 ± 10.2 vs. 22.6 ± 7.1%, for patients and controls, respectively; P = 0.20; girls: 28.2 ± 5.7 vs. 26.4 ± 7.7%; P = 0.91). Also, no differences were found regarding FFM% in boys (74.9 ± 10.2 vs. 77.4 ± 7.1%; P = 0.22) and girls (71.8 ± 5.6 vs. 73.5 ± 7.7%; P = 0.85). Conclusion: BIA adequately assesses body composition (FM%) in children with IBD and could advantageously replace DEXA, which is more expensive and less available. No differences in body composition were found between children with IBD and healthy controls.

CDK10/cyclin M is a protein kinase that controls ETS2 degradation and is deficient in STAR syndrome.

Cyclin-dependent kinases (CDKs) regulate a variety of fundamental cellular processes. CDK10 stands out as one of the last orphan CDKs for which no activating cyclin has been identified and no kinase activity revealed. Previous work has shown that CDK10 silencing increases ETS2 (v-ets erythroblastosis virus E26 oncogene homolog 2)-driven activation of the MAPK pathway, which confers tamoxifen resistance to breast cancer cells. The precise mechanisms by which CDK10 modulates ETS2 activity, and more generally the functions of CDK10, remain elusive. Here we demonstrate that CDK10 is a cyclin-dependent kinase by identifying cyclin M as an activating cyclin. Cyclin M, an orphan cyclin, is the product of FAM58A, whose mutations cause STAR syndrome, a human developmental anomaly whose features include toe syndactyly, telecanthus, and anogenital and renal malformations. We show that STAR syndrome-associated cyclin M mutants are unable to interact with CDK10. Cyclin M silencing phenocopies CDK10 silencing in increasing c-Raf and in conferring tamoxifen resistance to breast cancer cells. CDK10/cyclin M phosphorylates ETS2 in vitro, and in cells it positively controls ETS2 degradation by the proteasome. ETS2 protein levels are increased in cells derived from a STAR patient, and this increase is attributable to decreased cyclin M levels. Altogether, our results reveal an additional regulatory mechanism for ETS2, which plays key roles in cancer and development. They also shed light on the molecular mechanisms underlying STAR syndrome.