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A group of scientists interested in history of science and fascinated by the figure of Nicolaus Steno (1638-1686) gathered in Florence for the 350th anniversary of the publication of his De solido intra solidum naturaliter contento prodromus dissertationis. A public conference held at Palazzo Fenzi on 16 October 2019 and a geological fieldtrip on the following day were occasions to discuss different points of view on the last published work of the Danish natural philosopher, dedicated to "solids naturally enclosed in other solids" (De solido intra solidum naturaliter contento, or De solido in short). The title of the gathering, "Galilean foundation for a solid earth", emphasized the philosophical context that Steno found in Florence, where in 1666-1668 he established tight human and philosophical bonds with renowned Italian disciples of Galileo Galilei and members of the Accademia del Cimento. For participants to the 2019 gathering, the Museum of Natural History of the University of Florence, hosting some of Steno's geological specimens, and the region of Tuscany itself, formed the perfect location to discuss the phenomena that Steno had observed from 1666-1668, the motivations for his research, the methodology of his discovery and, generally stated, the European scientific context which informed his inquiry. Some of the talks given in that meeting are included within this volume, kindly hosted by Substantia, International Journal of the History of Chemistry published by the Florence University Press. In addition some of the invited speakers who were unable to attend, also contributed a paper to this publication. The collection is about earth science in the early modern period, when the study of minerals, rocks, and the fossilized remains of living things did not yet form a distinct path to knowledge about earth history, but was an integral part of the wider "philosophy of nature".

Purpose To investigate the efficacy of a micro-emulsion (ME) lipid layer artificial tear in improving tolerance of immunomodulator eye drops for the treatment of dry eye disease. Patients and Methods A total of 33 patients with previously diagnosed dry eye disease were given the micro-emulsion lipid layer artificial tear in conjunction with either lifitegrast or cyclosporine. Patients were queried on their tolerance of the regimen by reporting VAS scores before starting the adjunctive eye drop, immediately after starting, and 2-3 weeks later. Tolerance was statistically compared over time and with respect to previous medication compliance, timing of the adjunctive eye drop, age, gender, and ethnicity. Results Across all patients, the VAS pre-treatment score (6.8 ± 0.6) was significantly higher than both the VAS 1-day post ME lipid tear instillation time point (3.0 ± 0.7) (post hoc Bonferroni, p < 0.001) and the VAS 2-3-week post instillation time point (1.7 ± 0.7) (post hoc Bonferroni, p < 0.001), with the mean VAS score improving over time (post hoc Bonferroni, p < 0.028). Average VAS scores did not vary with respect to specific medical therapy or the timing of instillation of this artificial tear. Both the "at-risk" and "conversion" groups independently had significant improvements at both 1-day and 2-3-week time points compared to baseline. Conclusion The micro-emulsion lipid layer artificial tear was effective as an adjunctive eye drop to improve tolerance of lifitegrast and cyclosporine for patients with dry eye disease who were at risk of failing or had previously failed immunomodulatory therapy.

Purpose Bacterial burden on the eyelid margin and within meibomian glands was evaluated for influence on specific ocular surface disease (OSD) markers across the meibomian gland dysfunction (MGD) spectrum. Methods In this prospective, observational, single-center study, 40 patients were divided into 4 equal groups of 10 that encompassed increasingly worse MGD/OSD categories. All patients answered the standard Ocular Surface Disease Index questionnaire, and underwent tear osmolarity testing (TOT), Schirmer 1, matrix metalloproteinase 9 (MMP-9) testing, meibography, and lissamine green staining. Cultures of eyelid margins and meibomian gland secretions were directly plated on blood, chocolate, and Sabouraud agar; smears were sent for gram and Papinicolau evaluation. Results Mean patient age was 55.25±17.22 years; there were 10 males and 30 females. TOT and MMP-9 testing were similar across groups. Culture positivity was 62.5% for right eyes, 70% for left eyes, and was not statistically different across groups (for both eyelid margin and meibomian glands). The majority of cultures were positive for coagulase-negative staphylococcus (CNS). Conclusion This study is in concordance with others, citing the predominance of CNS within the biofilm of both "normal" and clinically significant MGD/OSD patients. Our study exemplifies that symptoms of OSD do not necessarily correlate with degree of clinical exam findings, nor culture positivity. These results argue that bacterial burden should be reconsidered as a direct risk factor and treatment target for MGD/OSD patients.

OBJECTIVE. The purpose of this study was to determine whether a single, uniform normalized iodine threshold reduces variability and enables reliable differentiation between vascular and nonvascular renal lesions independent of the dual-energy CT (DECT) platform used. MATERIALS AND METHODS. In this retrospective, HIPAA-compliant, institutional review board-approved study, 247 patients (156 men, 91 women; mean age ± SD, 67 ± 12 years old) with 263 renal lesions (193 nonvascular, 70 vascular) underwent unenhanced single- energy and contrast-enhanced DECT scans. One hundred and six nonvascular and 38 vascular lesions were scanned on two dual-source DECT (dsDECT) scanners, and 87 nonvascular and 32 vascular lesions were scanned on two rapid-kilovoltage-switching single-source DECT (rsDECT) scanners. Optimal absolute and normalized (to aorta) lesion iodine thresholds were determined for each platform type and for the entire cohort combined. RESULTS. Mean optimal absolute discriminant thresholds were 1.3 mg I/mL (95% CI, 1.2-1.9 mg I/mL), 1.6 mg I/mL (95% CI, 0.9-1.5 mg I/mL), and 1.5 mg I/mL (95% CI, 1.4- 1.7 mg I/mL) for dsDECT, rsDECT, and combined cohorts, respectively. Optimal normalized discriminant thresholds were 0.3 mg I/mL (95% CI, 0.2-0.4 mg I/mL) for both the dsDECT and rsDECT cohorts, and 0.3 mg I/mL (0.3-0.4 mg I/mL) for the combined cohort. The AUC, sensitivity, and specificity for the combined optimal normalized discriminant threshold of 0.3 mg I/mL was 0.96 (95% CI, 0.92-1.00), 0.93 (0.84-0.97), and 0.95 (0.91-0.98), respectively. Normalization resulted in decreased variability and better lesion separation (effect size, 1.77 vs 1.69, p < 0.0001). CONCLUSION. The optimal absolute discriminant threshold for evaluating renal lesions varies depending on the type of DECT platform, though this difference is not statistically significant. Variation can be reduced with a better separation of vascular and nonvascular lesions by normalizing iodine quantification to the aorta.

Spring runoff often comprises the majority of annual discharge and riverine phosphorus (P) export due to sustained high flow, and the magnitude of spring runoff can be a strong predictor of receiving water summer harmful algal bloom severity. Yet the loading of reactive forms of P during this time period remains poorly-characterized in time, space and geochemical partitioning. Here, we explore the hypothesis that riverine dissolved and suspended sediment P loads during spring runoff have a particularly high proportion of potentially reactive species due to unique hydrologic pathways and P association with iron (Fe).The concentration, distribution and temporal dynamics of dissolved P (DP), dissolved and colloidal Fe, and redox sensitive suspended sediment P (RSP) and Fe during spring runoff and summer storms were compared in forested and agricultural catchments of the same watershed. The dominant carrier of RSP was Fe (oxy)hydroxides across land cover and season, but Fe (oxy)hydroxide particles and colloids in agricultural catchments were strongly enriched in RSP and DP during spring runoff and summer storms, particularly at the onset of snowmelt. In 2014, 83% of DP and 74% of RSP were delivered to Missisquoi Bay during spring runoff. Suspended sediment was significantly more redox sensitive than typically input to limnological models, suggesting that the reactivity of this load may be systematically underestimated. Changes in the timing, provenance and severity of spring runoff associated with climate or land cover change will have dramatic impacts on total riverine P loads and their potential reactivity in receiving water ecosystems.

Objectives Nasal reconstruction is limited by the availability of autologous cartilage. The aim is to investigate an adhesive biomaterial for tissue engineering of nasal cartilage by evaluating mechanical properties of hydrogels made of fibrin crosslinked with genipin as compared to native tissue. Methods Hydrogels of fibrin, fibrin-genipin, and fibrin-genipin with extracellular matrix (ECM) particles were created and evaluated with mechanical testing to determine compression, tensile, and shear properties. Rabbit nasal septal cartilage was harvested and tested in these modalities for comparison. Transmission electron microscopy characterized hydrogel structure. Results Fibrin-genipin gels had higher compressive, tensile, and shear moduli compared to fibrin alone or fibrin-genipin with ECM. However, all hydrogel formulations had lower moduli than the rabbit nasal septal cartilage. Electron microscopy showed genipin crosslinking increased structural density of the hydrogel and that cartilage ECM created larger structural features with lower crosslinking density. Conclusion The addition of genipin significantly improved mechanical properties of fibrin hydrogels by increasing the compressive, tensile, and shear moduli. The addition of cartilage ECM, which can add native structure and composition, resulted in decreased moduli values. Fibrin-genipin is a bioactive and biomechanically stable hydrogel that may offer promise as a scaffold for cartilage tissue engineering in nasal reconstruction, yet further augmentation is required to match material properties of native nasal cartilage.

International audience; Unprecedented ground-based and aircraft measurements in June-July 2016 in southern West Africa characterize atmospheric composition and dynamics, low-level cloud properties, the diurnal cycle, and air pollution impacts on health.The EU-funded project DACCIWA (Dynamics-Aerosol-Chemistry-Cloud Interactions in West Africa) investigates the relationship between weather, climate, and air pollution in southern West Africa, an area with rapid population growth, urbanisation, and increase in anthropogenic aerosol emissions. The air over this region contains a unique mixture of natural and anthropogenic gases, liquid droplets and particles, emitted in an environment, in which multi-layer clouds frequently form. These exert a large influence on the local weather and climate, mainly due to their impact on radiation, the surface energy balance, and thus the diurnal cycle of the atmospheric boundary layer.In June and July 2016, DACCIWA organized a major international field campaign in Ivory Coast, Ghana, Togo, Benin, and Nigeria. Three supersites in Kumasi, Savè, and Ile-Ife conducted permanent measurements and 15 Intensive observation periods. Three European aircraft together flew 50 research flights between 27 June and 16 July 2016 for a total of 155 hours. DACCIWA scientists launched weather balloons several times a day across the region (772 in total), measured urban emissions, and evaluated health data. The main objective was to build robust statistics of atmospheric composition, dynamics, and low-level cloud properties in various chemical landscapes to investigate their mutual interactions.This article presents an overview of the DACCIWA field campaign activities as well as some first research highlights. The rich data obtained during the campaign will be made available to the scientific community and help to advance scientific understanding, modeling, and monitoring the atmosphere over southern West Africa.

Aerosol chemical composition was found to influence nighttime atmospheric chemistry during a series of airborne measurements in northwestern Europe in summer conditions, which has implications for regional air quality and climate. The uptake of dinitrogen pentoxide, γ (N2O5), to particle surfaces was found to be modulated by the amount of water content and ammonium nitrate present in the aerosol. The conditions prevalent in this study suggest that the net uptake rate of N2O5 to atmospheric aerosols was relatively efficient compared to previous studies, with γ (N2O5) values in the range 0.01–0.03. This is likely a consequence of the elevated relative humidity in the region, which promotes greater aerosol water content. Increased nitrate concentrations relative to particulate water were found to suppress N2O5 uptake. The results presented here contrast with previous ambient studies of N2O5 uptake, which have generally taken place in low-nitrate environments in the USA. Comparison of the N2O5 uptake derived from the measurements with a parameterised scheme that is based on the ratio of particulate water to nitrate yielded reasonably good agreement in terms of the magnitude and variation in uptake, provided the effect of chloride was neglected. An additional suppression of the parameterised uptake is likely required to fully capture the variation in N2O5 uptake, which could be achieved via the known suppression by organic aerosol. However, existing parameterisations representing the suppression by organic aerosol were unable to fully represent the variation in N2O5 uptake. These results provide important ambient measurement constraint on our ability to predict N2O5 uptake in regional and global aerosol models. N2O5 uptake is a potentially important source of nitrate aerosol and a sink of the nitrate radical, which is the main nocturnal oxidant in the atmosphere. The results further highlight the importance of ammonium nitrate in northwestern Europe as a key component of atmospheric composition in the region.

Dry eye disease (DED) is a common ocular disorder that can have a substantial burden on quality of life and daily activities. Lifitegrast ophthalmic solution 5.0% is the first medication approved in the US for the treatment of the signs and symptoms of DED. The aim of this article is to summarize the preclinical and clinical data on lifitegrast and discuss how lifitegrast may fit into the current treatment landscape for DED. Areas covered: A literature search of published preclinical and clinical data was conducted to review the chemistry, pharmacodynamics, pharmacokinetics, and clinical efficacy/safety of lifitegrast. The impact that lifitegrast may have on DED treatment practices is also discussed. Expert opinion: The introduction of lifitegrast provides a potentially important additional option for eye care professionals treating DED. In clinical trials conducted in adults with DED, lifitegrast ophthalmic solution 5.0% improved both signs and symptoms of DED. Of note, in 2 phase 3 trials, symptom improvements were observed as early as 2 weeks, which may be explained by lifitegrast's unique mechanism of action of blocking a specific signaling pathway in inflammation. Future research should include evaluation of whether lifitegrast can be used in combination with other DED treatments.

Hemostasis is a major concern during the perioperative period. Changes in platelet aggregation and coagulation factors may contribute to the delicate balance between thrombosis and bleeding. We sought to better understand perioperative hemostasis by investigating the changes in platelet aggregation and coagulation factors during the perioperative period. We performed a prospective cohort analysis of 70 subjects undergoing non-emergent orthopedic surgery of the knee (n = 28), hip (n = 35), or spine (n = 7) between August 2011 and November 2011. Plasma was collected preoperatively (T1), 1-h intraoperatively (T2), 1-h (T3), 24-h (T4) and 48-h (T5) postoperatively. Platelet function testing was performed using whole blood impedance aggregometry. Coagulation assays were performed for factor VII, factor VIII, von Willebrand Factor (vWF), and fibrinogen. Of the 70 patients, mean age was 64.1 ± 9.8 years, 61 % were female, and 74 % were Caucasian. Platelet activity decreased until 1 h postoperatively and then significantly increased above baseline at 24- and 48-h postoperatively. Compared to baseline, coagulation factors decreased intraoperatively. Factor VII activity continued to decrease, while FVIII, vWF, and fibrinogen all increased above baseline postoperatively. The results of our study indicate significant changes in platelet activity and coagulation factors during the perioperative period. Both platelet activity and markers of coagulation decrease during the intraoperative period and then some increase postoperatively. These changes may contribute to the hypercoagulabity and/or bleeding risk that occurs in the perioperative period. Future prospective studies aimed at correlating hemostatic changes with perioperative outcomes are warranted.

A new class of osteoconductive and osteoinductive combination biomaterials composed of calcium phosphate cement (CPC), demineralized bone matrix (DBM) and a water-soluble viscosity modifier were prepared and characterized in-vitro and in-vivo. In previous studies, a range of combinations formulations were tested in order to compare their performance characteristic. In-vitro characterization results show that the mechanical strength is decreased when the amount of DBM increases. However, DBM does not affect the CPC’s ability to set hard and convert to nanocrystalline apatitic calcium phosphate, which shares the chemical structure of natural bone as seen in x-ray diffraction. It is known that the DBM alone is osteoinductive. In-vivo osteoinductivity testing of the formulations in an intramuscular, athymic rat model demonstrated that the combination material is also osteoinductive. Two formulations were chosen for in-vivo efficacy testing based on the results of in-vitro and in-vivo characterization. These formulations were studied using rabbit critical-sized femoral core defect model. The formulations were composed of DBM with particle sizes of 250 to 710 μm, carboxymethyl-cellulose (CMC) as the viscosity modifier and weight percent compositions of 50% DBM/ 45% CPC/ 5% CMC and 60% DBM/ 30% CPC/ 10% CMC. Bone integration and healing was graded at 6, 12, and 24 weeks. The two formulations were compared to the gold standard autograft at 12 weeks and to an empty defect as the negative control at 24 weeks. Based on micro-computed topography (μCT), both formulations allowed for continuity of bone throughout the defect region at all time points. No differences in dense area fraction were seen between two formulations at 6 weeks (p = 0.8661). There was no significant statistical difference between the two formulations and autograft at 12 weeks (p = 0.2467). At 24 weeks, both formulations had significantly higher dense area fractions than empty controls (p = 0.0001). Histologically, the biology of the treatment areas appeared to have returned to normal by 24 weeks with CPC appearing to be the principal osteogenic inducer. In conclusion, these combinations of CPC and DBM offers significant advantages (handling, mechanical properties and osteoinductivity) over current DBM products and can be an effective alternative to autograft in healing of bone defects.

Alpha-bsm® is a first generation self-setting, injectable and moldable apatitic calcium phosphate cement (CPC) based on amorphous calcium phosphate (ACP). ACP was prepared using low temperature double decomposition technique, from a calcium solution (0.16 M), and phosphate solution (0.26 M) in a basic (pH~13) media. ACP was than stabilized using three crystal growth inhibitors (CO32-, Mg2+, P2O74-), freeze-dried, and heated (450 °C, 1h) to remove additional moisture and some inhibitors. Dicalcium phosphate dehydrate (DCPD) was also prepared using wet chemistry at room temperature from calcium and phosphate solution, respectively, 0.3 M and 0.15 M. ACP and DCPD powder were combined at a 1:1 ratio and ground to produce Alpha-bsm® bone cement. The cement is supplied as a powder and when mixed with an appropriate amount (0.8 ml/g) of physiological saline at room temperature, forms an injectable putty-like paste. The paste has a working time of about 45 minutes at room temperature, when stored in a moist environment. The setting reaction proceeds isothermically at body temperature (37°C) in less than 20 minutes, forming a hardened, porous (total porosity 50 to 60%), low crystalline (40% comparing with HA), apatitic calcium phosphate cement with a compressive strength range of 10 to 12 MPa. Extensive pre-clinical studies (rabbit radius critical sized defect, canine tibia osteotomy, sheep tibia, primate fibula fracture healing, and primate fibula critical size defect) demonstrate that Alpha-bsm® undergoes remodeling in conjunction with new bone formation. The next generation of Bone Substitute Materials (Beta-bsmTM and Gamma-bsm TM) are formulated based on the Alpha-bsm® chemistry but differ in powder processing (e.g. milling) technique. These materials are also self-setting, injectable and/or moldable apatitic calcium phosphate cements with improved handling and mechanical properties. The setting & hardening reaction of these new CPCs proceeds isothermically in less than 5 minutes at 37°C and once hardened demonstrate a compressive strength of 30 to 50 MPa. The final product (after full conversion) is a low crystalline (40% compared with Hydroxyapatite), calcium deficient (Ca/P atomic ratio = 1.45) carbonated apatite similar to the composition and structure of natural bone mineral (crystal size: length = 26 nm, width thickness = 8 nm). A desirable feature of these cements is their high surface chemistry (with specific surface area of about 180-200 m2/g) which is ideal for remodeling and controlled release of growth factors. A pilot rabbit critically sized femoral defect study comparing the three synthetic family products demonstrate that they share similar remodeling and resorption characteristics up to 52 weeks. Physico-chemical and mechanical performance of these next generation CPCs are favorable when compared with existing CPCs in the market, specifically material working time (at room temperature), cohesivity in a wet environment and fast setting & hardening rate (at body temperature).

Aboard the Huygens probe, which descended through Titan's atmosphere in January 2005, was the Surface Science Package (SSP), a set of 9 sensors, including a speed-of-sound sensor. We present the first detailed description of the SSP speed of sound measurements and report constraints on the methane content in Titan's lower atmosphere based on these measurements. After a careful calibration and subsequent Bayesian analysis, our measurements yield a most likely methane fraction in Titan's lower atmosphere of approximately 2% at 10 km, increasing to 3.5% at lower altitudes, based on a binary composition. Our data show that any large scale variation of methane within the lower 11 km of Titan's atmosphere is unlikely. Within experimental and theoretical uncertainties, our results are compatible with earlier estimates obtained from the GCMS experiment [Niemann, H.B., Atreya, S.K., Bauer, S.J., Carignan, G.R., Demick, J.E., Frost, R.L., Gautier, D., Haberman, J.A., Harpold, D.N., Hunten, D.M., Israel, G., Lunine, J.I., Kasprzak, W.T., Owen, T.C., Paulkovich, M., Raulin, F., Raaen, E., Way, S.H., 2005. Nature 438 (7069), 779–784].

We report on three patients with hepatitis C virus infection, severe mental illness, and substance use disorders that we treated successfully with interferon and ribavirin. This population has historically been refused such treatment on the grounds that they were unlikely to be adherent to treatment or would experience untoward psychiatric side effects. These case reports add support to the growing body of evidence that persons with severe mental illness and substance use disorders can be successfully treated for hepatitis C with careful monitoring and psychiatric oversight.

We present a method for inferring the relative molar abundance of constituents of a liquid mixture, in this case methane, ethane, nitrogen and argon, from a measurement of a set of physical properties of the mixture. This problem is of interest in the context of the Huygens Surface Science Package, SSP, equipped to measure several physical properties of a liquid in case of a liquid landing on Saturn's moon Titan. While previous models emphasized the possibility of verifying a certain model proposed by atmospheric composition and equations of state, we use an inverse approach to the problem, i.e. we will infer the liquid composition strictly from our measurements of density, refractive index, permittivity, thermal conductivity and speed of sound. Other a priori information can later be used to improve (or reject) the model obtained from these measurements.

Resume Les recepteurs hormonaux membranaires sont repartis en trois grandes familles : les recepteurs couples aux proteines G, les recepteurs tyrosine kinase et les recepteurs des cytokines. Chacune de ces familles est caracterisee par des proprietes structurales communes et des mecanismes de transduction du signal particuliers.

BACKGROUND: PLTs stored at 22 degrees C have the potential for bacterial contamination, a problem that could be reduced by 4 degrees C storage. Nevertheless, PLTs stored at 4 degrees C exhibit a significantly reduced life span. This study was performed to determine whether treatment of PLTs with thrombopoietin or cytochalasin B plus ethylene glycol bis(beta-aminoethyl ether) N,N'-tetraacetic acid (EGTA)-AM could prevent exponential loss of PLTs stored at 4 degrees C. STUDY DESIGN AND METHODS: Autologous baboon PLTs were stored at 22 or 4 degrees C. The 4 degrees C stored PLTs were treated with 1.5 ng per mL thrombopoietin, with 1 micro mol per L cytochalasin B, and 80 micromol per L EGTA-AM (cyto-EGTA) or not treated and labeled with (111)In-oxine to study their in vivo recovery and life span. PLT function was assessed by correction of an aspirin-induced prolonged bleeding time. Aggregation responses and morphology were also assessed. RESULTS: PLTs stored at 22 degrees C had normal in vivo recovery and linear survival. PLTs stored at 4 degrees C, whether or not they were treated with thrombopoietin, had normal recovery and exponential survival. Aggregation of cyto-EGTA-treated PLTs was similar for PLTs stored at 4 degrees C and fresh PLTs, but decreased in PLTs stored at 22 degrees C for 5 days. The addition of cyto-EGTA to PLTs before 4 degrees C storage inhibited morphologic changes that occurred in PLTs stored at 22 degrees C and cold-induced PLT clumping, but did not prevent exponential disappearance of the PLTs. CONCLUSION: Addition of thrombopoietin or cyto-chalasin B and EGTA-AM to PLTs before 4 degrees C storage did not prevent exponential loss of PLTs.

Sulfated motifs on heparan sulfate (HS) are involved in various extracellular processes from cell signaling to enzymatic regulation, but the structures of these motifs are obscure. We have developed a strategy to determine the structure of sulfotransferase recognition sites which constitute these motifs. Stable isotope is first introduced into specific sites on HS with HS sulfotransferases and the modified HS is then digested into oligosaccharides of differing sizes. The overlapping oligosaccharides containing the introduced stable isotope are identified by changes in the m/z profiles by mass spectrometry, and their relationships are elucidated. In this way, the HS structures in the vicinity of the sulfotransferase recognition site are quickly determined and groups on precursor structures of HS that direct the action of HS sulfotransferases are pinpointed.

Heparan sulfate, a cell surface bound glycosaminoglycan polysaccharide, has been implicated in numerous biological functions. Heparan sulfate molecules are highly complex and diverse, yet deceivingly look simple and similar, rendering structure--function correlation tedious. Current chromatographic and mass spectrometric techniques have limitations for analyzing glycosaminoglycan samples that are in low abundance and that are large in size, due to their highly acidic nature arising from a large number of sulfate and of carboxylate groups. A new methodology was developed using capillary ion-paired reverse-phase C18 HPLC directly coupled to ESI-TOF-MS to address the above issues. On the basis of HS disaccharide analysis, dibutylamine was found to be the best suited for HS analysis among many ion-pairing agents investigated. Next, analysis of oligosaccharides derived from heparosan, the precursor for heparan sulfate, was undertaken to demonstrate its greater applicability in a more complex structural analysis. The established chromatographic conditions enabled the characterization of heparosan oligosaccharides of sizes up to tetracontasaccharide with high resolution in a single run and were amenable to negative ion electrospray MS in which sodium adduction and fragmentation were avoided. To date, these are the largest nonsulfated HS precursor oligosaccharides to be characterized by LC/MS. Finally, the current methodology was applied to the characterization of the biologically important ATIII binding pentasaccharide and its precursors, which differ from each other by sulfation pattern and/or degree of sulfation. All of these pentasaccharides were well-resolved and characterized by the LC/MS system with (34)SO(4) as a mass spectral probe. This newly developed methodology facilitates the purification and rapid characterization of biologically significant HS oligosaccharides, and will thus expedite their synthesis. These findings should undoubtedly pave the way in deciphering multiple functional arrangements, ascribed to many biological activities, which are predictably embedded in a single large chaotic, yet well-organized HS polysaccharide chain. Development of newer techniques for HS oligosaccharide analysis is greatly needed in the postgenome era as attention shifts to the functional implications of proteins and carbohydrates in general and HS in particular.

On the eastern flank of the Juan de Fuca Ridge, reaction between upwelling basement fluid and sediment alters hydrothermal fluxes of Ca, SiO2(aq), SO4, PO4, NH4, and alkalinity. We used the Global Implicit Multicomponent Reactive Transport (GIMRT) code to model the processes occurring in the sediment column (diagenesis, sediment burial, fluid advection, and multicomponent diffusion) and to estimate net seafloor fluxes of solutes. Within the sediment section, the reactions controlling the concentrations of the solutes listed above are organic matter degradation via SO4 reduction, dissolution of amorphous silica, reductive dissolution of amorphous Fe(III)-(hydr)oxide, and precipitation of calcite, carbonate fluorapatite, and amorphous Fe(II)-sulfide. Rates of specific discharge estimated from pore-water Mg profiles are 2 to 3 mm/yr. At this site the basement hydrothermal system is a source of NH4, SiO2(aq), and Ca, and a sink of SO4, PO4, and alkalinity. Reaction within the sediment column increases the hydrothermal sources of NH4 and SiO2(aq), increases the hydrothermal sinks of SO4 and PO4, and decreases the hydrothermal source of Ca. Reaction within the sediment column has a spatially variable effect on the hydrothermal flux of alkalinity. Because the model we used was capable of simulating the observed pore-water chemistry by using mechanistic descriptions of the biogeochemical processes occurring in the sediment column, it could be used to examine the physical controls on hydrothermal fluxes of solutes in this setting. Two series of simulations in which we varied fluid flow rate (1 to 100 mm/yr) and sediment thickness (10 to 100 m) predict that given the reactions modeled in this study, the sediment section will contribute most significantly to fluxes of SO4 and NH4 at slow flow rates and intermediate sediment thickness and to fluxes of SiO2(aq) at slow flow rates and large sediment thickness. Reaction within the sediment section could approximately double the hydrothermal sink of PO4 over a range of flow rates and sediment thickness, and could slightly decrease (by ≤10%) the size of the hydrothermal source of Ca.

The composition of Jupiter and the primordial distribution of the heavy elements are determined by its formation history. As a result, in order to constrain the primordial internal structure of Jupiter the growth of the core and the deposition and settling of accreted planetesimals must be followed in detail. In this paper we determine the distribution of the heavy elements in proto-Jupiter and determine the mass and composition of the core. We find that while the outer envelope of proto-Jupiter is typically convective and has an homogeneous composition, the innermost regions have compositional gradients. In addition, the existence of heavy elements in the envelope leads to much higher internal temperatures (several times 10$^4$ K) than in the case of a hydrogen-helium envelope. The derived core mass depends on the actual definition of the core: if the core is defined as the region in which the heavy-element mass fraction is above some limit (say 0.5), then it can be much more massive ($\sim$ 15 M$_{\oplus}$) and more extended (10\% of the planet's radius) than in the case where the core is just the region with 100\% heavy elements. In the former case Jupiter's core also consists of hydrogen and helium. Our results should be taken into account when constructing internal structure models of Jupiter and when interpreting the upcoming data from the {\it Juno} (NASA) mission., Comment: 32 pages, 12 figures

An estimated 290,000 surgical site infections occur after a procedure in the United States annually, accounting for 22% of all healthcare associated infections [1]. Deep surgical site infections (SSI) after arthroplasty or spine fusion surgery complicate up to 2% of cases, and result in revision surgery and prolonged antibiotic use [2, 3]. The patient morbidity and healthcare system cost is tremendous, with an estimated $566 million spent annually in hospital treatment costs for arthroplasty SSI alone [4]. Staphylococcus aureus is a frequent and feared cause of these infections, given its unique pathogenicity and ability to adhere to prosthetic material [5, 6]. Studies indicate S. aureus colonization prior to surgery is a risk of subsequent infection, with the nasal mucosa serving as a reservoir for S. aureus colonization and a source of secondary transmission to other body sites [7, 8]. Prevention of SSI by treatment of S. aureus colonization with intranasal topical mupirocin has been studied. A short-term suppression rate of 83% after multiple doses of nasal mupirocin was achieved in one randomized, placebo-controlled trial of 891 S. aureus colonized patients, resulting in a statistically significant reduction of invasive S. aureus infection [9]. Several controlled trials suggest a reduction in SSI with the use of pre-operative topical antiseptics [10, 11]. When nasal mupirocin was combined with use of chlorhexidine soap in a randomized, double-blind, placebo-controlled trial including 808 S. aureus colonized surgical patients, a significant reduction in deep S. aureus SSI was realized [12]. To reduce the risk of SSI after arthroplasty and spine fusion surgery at our institution, we historically provided a prescription for brand mupirocin ointment specifically formulated for application on intranasal mucosal surfaces twice a day for the five days prior to surgery, and instructions for the use of chlorhexidine soap the evening before surgery. After implementation of this protocol, we conducted an anonymous patient survey to measure compliance. Although 94% of patients used the chlorhexidine soap, only 86% applied the mupirocin ointment and 8% of patients stated they found it hard or very hard to purchase the mupirocin due to cost [13]. The brand nasal mupirocin ointment specifically produced for application on intranasal mucosal surfaces is only formulation currently available; although generic mupirocin ointment for topical use on skin is available at less cost, application of this formulation on mucosal surfaces may cause irritation. Our survey results, plus reports of emerging mupirocin resistance, led us to search for alternatives [14–19]. Povidone-iodine solution is a broad-spectrum antiseptic suitable for suppression of S. aureus in nasal secretions [20]. In contrast to the application of nasal mupirocin antibiotic ointment to eradicate S. aureus in the nares before surgery, the application of povidone iodine is intended to transiently suppress S. aureus in the nares during surgery. Our hypothesis was a one-time application of nasal povidone iodine just prior to surgery would be as effective as twice daily applications of nasal mupirocin during the five days before surgery in preventing SSI, and provide a more convenient option for patients at lower cost.

Background: Neuraxial local anesthetics may have neurological complications thought to be due to neurotoxicity. A primary site of action of local anesthetics is the dorsal root ganglia (DRG) neuron. Physiologic differences have been noted between young and adult DRG neurons; hence, the authors examined whether there were any differences in lidocaine-induced changes in calcium and lidocaine toxicity in neonatal and adult rat DRG neurons. Methods: DRG neurons were cultured from postnatal day 7 (P7) and adult rats. Lidocaine-induced changes in cytosolic calcium were examined with the calcium indicator Fluo-4. Cells were incubated with varying concentrations of lidocaine and examined for viability using calcein AM and ethidium homodimer-1 staining. Live imaging of caspase-3/7 activation was performed after incubation with lidocaine. Results: The mean KCl-induced calcium transient was greater in P7 neurons (P < 0.05), and lidocaine significantly inhibited KCl-induced calcium responses in both ages (P < 0.05). Frequency distribution histograms of KCl-evoked calcium increases were more heterogeneous in P7 than in adult neurons. With lidocaine, KCl-induced calcium transients in both ages became more homogeneous but remained different between the groups. Interestingly, cell viability was decreased by lidocaine in a dose-dependent manner similarly in both ages. Lidocaine treatment also activated caspase-3/7 in a dose- and time-dependent manner similarly in both ages. Conclusions: Despite physiological differences in P7 and adult DRG neurons, lidocaine cytotoxicity is similar in P7 and adult DRG neurons in vitro. Differences in lidocaine- and KCl-evoked calcium responses suggest the similarity in lidocaine cytotoxicity involves other actions in addition to lidocaine-evoked effects on cytosolic calcium responses.

The authors report results from experiments on the evaporative chemical evolution of the two major types of natural waters present at Yucca Mountain, Nevada. The first is represented by J13 well water, a dilute Na–HCO3–CO3 water similar to saturated horizons in volcanic tuffs across the western United States. The second is represented by Ca–C1–SO4-rich pore water that has a higher dissolved ion content, from the unsaturated zone above the repository horizon. Data include anion and cation analysis and qualitative mineral identification for a series of open system experiments, with and without crushed tuff present, conducted at sub-boiling temperatures (75–85°C). This work is motivated by a need to characterize the chemistry of concentrated aqueous films that might form on engineered components at the potential high-level, nuclear-waste repository at Yucca Mountain, Nevada and lead to electrochemical corrosion. The experiments indicate the evolution of high pH, Na–HCO3–CO3 brines from J13-like waters and the evolution of near-neutral pH brines from the pore water compositions. The minerals recovered after complete evaporation of the J13 water alone experiments include amorphous silica, aragonite, calcite, halite, niter, thermonatrite and, possibly, gypsum, anhydrite and hectorite. Tachyhydrite and gypsum were observed to form in equivalent experiments with pore waters from the unsaturated zone.

This study investigates fluid absorption behavior of an unirradiated and a heavily crosslinked ultra-high molecular weight polyethylene (UHMWPE) exposed to bovine serum with pressures varying from 0.1 to 10 MPa. The time-response of serum absorption is biphasic, with an initial “soak-in” phase (

A concept has been developed for the production of photovoltaic energy collection systems on the Moon by vacuum deposition directly onto the lunar surface. Using this technique it would be possible to quickly install quite large capacities of power on the Moon at relatively low cost. Most of the material required for photovoltaic devices is readily available on the Moon, and taking production machinery to the Moon can further lower the cost of power if the machinery can be operated for long periods of time. Low-cost energy on the Moon could enable a wide range of activities including support of robust human outposts, production of propellants for use on the Moon or for export, production of a wide range of other materials for use in space, and beaming of energy from the Moon to space and to Earth. A proposed strategy by which lunar power can be developed in the near future is closely connected to the establishment of a human outpost on the Moon.

Results from an inorganic geochemical modeling study support a scenario in which low- temperature aqueous alteration of an anhydrous CM asteroidal parent body and melt water from H20 and CO;! ices produces the altered assemblage observed in CM carbonaceous chondrites (chrysotile, greenalite, tochilinite, cronstedtite and minor calcite and magnetite). We consider a range of possible precursor mineral assemblages, varying with respect to the Fe-oxidation state of the initial anhydrous phases. The aqueous solutions produced by this alteration are generally strongly basic and reducing and a large quantity of H2, and possible CH4, gas can be released during aqueous alteration.

SummaryThe risk of venous thrombosis is increased in individuals who carry specific genetic abnormalities in blood coagulation proteins. Among Caucasians, the prothrombin G20210A and factor V Arg506Gln (FV R506Q) mutations are the most prevalent defects identified to date. We evaluated their influence on markers of coagulation activation among participants in the Second Northwick Park Heart Study, which recruited healthy men (aged 50–61 years) from nine general medical practices in England and Wales. They were free of clinical vascular disease and malignancy at the time of recruitment. Genotypes for the two mutations were analyzed using microplate array diagonal gel electrophoresis, and coagulation markers (factor XIIa; activation peptides of factor IX, factor X, and prothrombin; fibrinopeptide A) were measured by immunoassay. Factor VII coagulant activity and factor VIIa levels were determined by a functional clotting assay. Among 1548 men genotyped for both mutations, 28 (1.8%) and 52 (3.4%) were heterozygous for prothrombin G20210A and FV R506Q, respectively. The only coagulation marker that was significantly associated with the two mutations was prothrombin activation fragment F1+2 [mean ± SD, 0.88 ± 0.32 nmol/L in men with prothrombin G20210A (p = 0.002) and 0.89 ± 0.30 in men with FV R506Q (p = 0.0001) versus 0.72 ± 0.24 among non-carriers for either mutation]. This data provides conclusive evidence that heterozygosity for the prothrombin G20210A as well as the FV R506Q mutations in the general population leads to an increased rate of prothrombin activation in vivo.

3-O-Sulfation of glucosamine by heparan sulfate D-glucosaminyl 3-O-sulfotransferase (3-OST-1) is the key modification in anticoagulant heparan sulfate synthesis. However, the heparan sulfates modified by 3-OST-2 and 3-OST-3A, isoforms of 3-OST-1, do not have anticoagulant activity, although these isoforms transfer sulfate to the 3-OH position of glucosamine residues. In this study, we characterize the substrate specificity of purified 3-OST-3A at the tetrasaccharide level. The 3-OST-3A enzyme was purified from Sf9 cells infected with recombinant baculovirus containing 3-OST-3A cDNA. Two 3-OST-3A-modified tetrasaccharides were purified from the 3-O-(35)S-sulfated heparan sulfate that was digested by heparin lyases. These tetrasaccharides were analyzed using nitrous acid and enzymatic degradation combined with matrix-assisted laser desorption/ionization-mass spectrometry. Two novel tetrasaccharides were discovered with proposed structures of DeltaUA2S-GlcNS-IdoUA2S-[(35)S]GlcNH(2)3S and DeltaUA2S-GlcNS-IdoUA2S-[3-(35)S]GlcNH(2)3S6S . The results demonstrate that 3-OST-3A sulfates N-unsubstituted glucosamine residues, and the 3-OST-3A modification sites are probably located in defined oligosaccharide sequences. Our study suggests that oligosaccharides with N-unsubstituted glucosamine are precursors for sulfation by 3-OST-3A. The intriguing linkage between N-unsubstituted glucosamine and the 3-O-sulfation by 3-OST-3A may provide a clue to the potential biological functions of 3-OST-3A-modified heparan sulfate.

To determine the role played by syndecan-4 cytoplasmic domain in the mediation of basic fibroblast growth factor (bFGF) signaling, immortalized human cells (ECV) were used to generate cell lines expressing constructs encoding full-length sequences for syndecan-4 (S4), syndecan-1 (S1), glypican-1 (G1), or chimeric proteins consisting of the ectoplasmic domain of glypican-1 linked to the transmembrane/cytoplasmic domain of syndecan-4 (G1-S4c) and the ectoplasmic domain of syndecan-4 linked to the glypican-1 glycosylphosphatidylinositol (GPI) anchor sequence (S4-GPI). Vector-transduced cells (VC) were used as controls. Expression of all these proteoglycans (except for the vector control) significantly increased cell-associated heparan sulfate mass and the number of low affinity bFGF-binding sites. However, in low serum medium, the addition of bFGF stimulated growth and migration of cells expressing S4 and G1-S4c constructs but not G1, S1, S4-GPI, or VC cells. Similar results were obtained using Matrigel growth assays. Mutations of heparan sulfate attachment sites on S4 construct abolished syndecan-4-dependent augmentation of bFGF responses. We conclude that cytoplasmic tail of syndecan-4 plays an important role in bFGF-mediated signal transduction.

3-O-Sulfated glucosaminyl residues are rare constituents of heparan sulfate and are essential for the activity of anticoagulant heparan sulfate. Cellular production of the critical active structure is controlled by the rate-limiting enzyme, heparan sulfate d-glucosaminyl 3-O-sulfotransferase-1 (3-OST-1) (EC 2.8.2.23). We have probed the expressed sequence tag data base with the carboxyl-terminal sulfotransferase domain of 3-OST-1 to reveal three novel, incomplete human cDNAs. These were utilized in library screens to isolate full-length cDNAs. Clones corresponding to predominant transcripts were obtained for the 367-, 406-, and 390-amino acid enzymes 3-OST-2, 3-OST-3A, and 3-OST-3B, respectively. These type II integral membrane proteins are comprised of a divergent amino-terminal region and a very homologous carboxyl-terminal sulfotransferase domain of ∼260 residues. Also recovered were partial length clones for 3-OST-4. Expression of the full-length enzymes confirms the 3-O-sulfation of specific glucosaminyl residues within heparan sulfate (Liu, J., Shworak, N. W., Sinay, P., Schwartz, J. J. Zhang, L., Fritze, L. M. S., and Rosenberg, R. D. (1999) J. Biol. Chem. 274, 5185–5192). Southern analyses suggest the human 3OST1, 3OST2, and 3OST4 genes, and the corresponding mouse isologs, are single copy. However, 3OST3A and 3OST3B genes are each duplicated in humans and show at least one copy each in mice. Intriguingly, the entire sulfotransferase domain sequence of the 3-OST-3B cDNA (774 base pairs) was 99.2% identical to the same region of 3-OST-3A. Together, these data argue that the structure of this functionally important region is actively maintained by gene conversion between 3OST3A and 3OST3B loci. Interspecific mouse back-cross analysis identified the loci for mouse3Ost genes and syntenic assignments of corresponding human isologs were confirmed by the identification of mapped sequence-tagged site markers. Northern blot analyses indicate brain exclusive and brain predominant expression of 3-OST-4 and 3-OST-2 transcripts, respectively; whereas, 3-OST-3A and 3-OST-3Bisoforms show widespread expression of multiple transcripts. The reiteration and conservation of the 3-OST sulfotransferase domain suggest that this structure is a self-contained functional unit. Moreover, the extensive number of 3OST genes with diverse expression patterns of multiple transcripts suggests that the novel 3-OST enzymes, like 3-OST-1, regulate important biologic properties of heparan sulfate proteoglycans.

The 3-O-sulfation of glucosamine residues is an important modification during the biosynthesis of heparan sulfate (HS). Our previous studies have led us to purify and molecularly clone the heparan sulfate d-glucosaminyl 3-O-sulfotransferase (3-OST-1), which is the key enzyme converting nonanticoagulant heparan sulfate (HSinact) to anticoagulant heparan sulfate (HSact). In this study, we expressed and characterized the full-length cDNAs of 3-OST-1 homologous genes, designated as 3-OST-2, 3-OST-3A, and 3-OST-3B as described in the accompanying paper (Shworak, N. W., Liu, J., Petros, L. M., Zhang, L., Kobayashi, M., Copeland, N. G., Jenkins, N. A., and Rosenberg, R. D. (1999)J. Biol. Chem. 274, 5170–5184). All these cDNAs were successfully expressed in COS-7 cells, and heparan sulfate sulfotransferase activities were found in the cell extracts. We demonstrated that 3-OST-2, 3-OST-3A, and 3-OST-3B are heparan sulfate d-glucosaminyl 3-O-sulfotransferases because the enzymes transfer sulfate from adenosine 3′-phosphophate 5′-phospho-[35S]sulfate ([35S]PAPS) to the 3-OH position of glucosamine. 3-OST-3A and 3-OST-3B sulfate an identical disaccharide. HSact conversion activity in the cell extract transfected by 3-OST-1 was shown to be 300-fold greater than that in the cell extracts transfected by 3-OST-2 and 3-OST-3A, suggesting that 3-OST-2 and 3-OST-3A do not make HSact. The results of the disaccharide analysis of the nitrous acid-degraded [35S]HS suggested that 3-OST-2 transfers sulfate to GlcA2S-GlcNS and IdoA2S-GlcNS; 3-OST-3A transfers sulfate to IdoA2S-GlcNS. Our results demonstrate that the 3-O-sulfation of glucosamine is generated by different isoforms depending on the saccharide structures around the modified glucosamine residue. This discovery has provided evidence for a new cellular mechanism for generating a defined saccharide sequence in structurally complex HS polysaccharide.

Retinoic acid (RA) and dibutyryl cAMP plus theophilline (CT) trigger F9 cells to differentiate into parietal endoderm. The differentiation induces a 9-fold increase in total heparan sulfate (HStotal) biosynthesis and a 170-fold increase in anticoagulantly active HS (HSact) biosynthesis. Measurement of 3-O-sulfotransferase-1 mRNA and enzymatic activity demonstrated an increase of over 100-fold whereas determination of N-, 2-O, and 6-O-sulfotransferase enzymatic activities showed elevations of 2-, 3. 5-, and 3.7-fold, respectively. HSact precursor pool measurements reveal that 30% of control F9 HStotal can be converted into HSact while only an additional 10% of RACT F9 HStotal can be transformed into HSact. Disaccharide analysis of metabolic labeled HS indicated that 32% 3-O-sulfate containing disaccharides, i.e. GlcA-anManR3S and GlcA-anManR3S6S, are present in HSact and 68% GlcA-anManR3S and GlcA-anManR3S6S are found in anticoagulantly inactive HS (HSinact). By using adenosine 3'-phosphate 5'-phosphosulfate and purified 3-O-sulfotransferase-1, 30% of 3-O-sulfation occurs in HSact and 70% of 3-O-sulfation occurs in HSinact. The similar ratio of 3-O-sulfate distribution in HSact versus HSinact suggests that HSact production in the F9 system is determined by the abundance of 3-O-sulfotransferase-1 as well as the size of the HSact precursor pool. Extensively 3-O-sulfated HSinact may play an important functional role under in vivo conditions within the murine placenta.

The diversity of cellular and tissue functions within organs requires that local communication circuits control distinct populations of cells. Recently, we reported that cardiac myocytes regulate the expression of both von Willebrand factor (vWF) and a transgene with elements of the vWF promoter in a subpopulation of cardiac microvascular endothelial cells (J. Cell Biol. 138:1117). The present study explores this communication. Histological examination of the cardiac microvasculature revealed colocalization of the vWF transgene with the PDGF alpha-receptor. Transcript analysis demonstrated that in vitro cardiac microvascular endothelial cells constitutively express PDGF-A, but not B. Cardiac myocytes induced endothelial expression of PDGF-B, resulting in PDGF-AB. Protein measurement and transcript analysis revealed that PDGF-AB, but not PDGF-AA, induced endothelial expression of vWF and its transgene. Antibody neutralization of PDGF-AB blocked the myocyte-mediated induction. Immunostaining demonstrated that vWF induction is confined to PDGF alpha-receptor-positive endothelial cells. Similar experiments revealed that the PDGF-AB/alpha-receptor communication also induces expression of vascular endothelial growth factor and Flk-1, critical components of angiogenesis. The existence of this communication pathway was confirmed in vivo. Injection of PDGF-AB neutralizing antibody into the amniotic fluid surrounding murine embryos extinguished expression of the transgene. In summary, these studies suggest that environmental induction of PDGF-AB/alpha-receptor interaction is central to the regulation of cardiac microvascular endothelial cell hemostatic and angiogenic activity.

During rodent fetal development, maternal lipoproteins can be sources of cholesterol for the membrane synthesis required for tissue growth in the developing embryo and for steroid hormone production in the extraembryonic tissues. Although the mechanisms underlying the maternal–fetal lipoprotein cholesterol transport system are not well defined, the placenta and yolk sac seem to play major roles in this process, serving as functionally active interfaces between maternal circulation and the embryo. In rodents, the principal cholesterol transporter in the plasma is HDL, and the HDL receptor SR-BI is a physiologically important mediator of cholesterol uptake in adult liver and steroidogenic tissues. To begin to investigate SR-BI's role in maternal cholesterol uptake by the fetus, we used immunofluorescence microscopy to determine the pattern of SR-BI expression during murine embryogenesis. At day E7.5 in gestation, there was significant SR-BI expression in endothelial cells of the decidua, but little in intraembryonic and extraembryonic tissues. By day E8.5, there was a dramatic increase in SR-BI expression in the trophoblast cells which surround the developing embryo. Beginning at day E10, SR-BI was expressed in both the placenta and yolk sac. The expression in these extraembryonic tissues was correlated with significant uptake of fluorescent dye by the yolk sac visceral endodermal cells from DiI-labeled HDL injected into pregnant mice. Within the embryo proper, SR-BI expression appeared by day E14.5 at high levels in the adrenal gland. SR-BI expression was not detected in the embryonic liver through day E17.5 of gestation; however, it could be observed in neonatal livers. These findings suggest that SR-BI may play a role in the rodent maternal–fetal lipoprotein cholesterol transport system, supplying HDL cholesterol for either membrane or steroid hormone synthesis, or both.—Hatzopoulos, A. K., A. Rigotti, R. D. Rosenberg, and M. Krieger. Temporal and spatial pattern of expression of the HDL receptor SR-BI during murine embryogenesis.

本論で我々は、心臓血管システムの三つの重要な機能分子-線維芽細胞増殖因子 (FGFs)、リポタンパク質リパーゼ、アンチトロンビン (AT)、の作用を介するHSPGの役割を紹介する。HSPGは、FGFレセプターを介するFGFのシグナル伝達を調節し、リポタンパク質リパーゼを重要な作用点に局在化させ、ATの抗凝固活性を直接刺激する。これらの多様な作用機構は、すべてそれぞれの機能分子とHS鎖の糖配列の間の高く特異的な相互作用によるということができよう。これらの糖配列は、主としてヘパラン硫酸鎖の中のN-、2-O-、6-O-、3-O-硫酸基の適切な配置により決定されている。すなわち、HSPGの生物学的機能は、HSの精巧な構造を決める生合成機構により制御されている。特に内皮細胞が作るAT結合部位は、3-O-スルホトランスフェラーゼ-1の律速活性により制御されている。知られているHSスルホトランスフェラーゼの構造と機能特性の比較から、異なる前駆体/生成物相関関係を持つ複数の異性体が存在し、少なくともその一部は順序の定まった生化学的経路の中で機能する。これらの観察から、我々は精巧なHS配列の合成制御のモデルを記述したい。

SummaryThe effects of chronic cigarette smoking on the coagulation system were examined in 2964 men aged 50 to 61 years and clinically free of cardiovascular disease. Factor VII activity (VIIc), factor VII antigen (VIIag), prothrombin fragment 1+2 (F1.2), fibrinopeptide A (FPA) and fibrinogen were measured in all participants, and activated factor VII (VIIa), factor IX activation peptide (IX pep) and factor X activation peptide (X pep) in a large sub-sample. The levels of all indices except FPA differed significantly between non-smokers, ex-smokers and current smokers. After adjustment for other conventional cardiovascular risk factors, mean VIIc was raised slightly by 3% in ex-smokers and current smokers as compared with non-smokers, owing to increases in VIIa and VIIag. Plasma IX pep, X pep, F1.2 and fibrinogen concentration were highest in current smokers, intermediate in ex-smokers and lowest in non-smokers. These findings accord with the increased risk of arterial thrombosis in smokers.

The cellular rate of anticoagulant heparan sulfate proteoglycan (HSPGact) generation is determined by the level of a kinetically limiting microsomal activity, HSact conversion activity, which is predominantly composed of the long sought heparan sulfate D-glucosaminyl 3-O-sulfotransferase (3-OST) (Shworak, N. W., Fritze, L. M. S., Liu, J., Butler, L. D., and Rosenberg, R. D. (1996) J. Biol. Chem. 271, 27063-27071; Liu, J., Shworak, N. W., Fritze, L. M. S., Edelberg, J. M., and Rosenberg, R. D. (1996) J. Biol. Chem. 271, 27072-27082). Mouse 3-OST cDNAs were isolated by proteolyzing the purified enzyme with Lys-C, sequencing the resultant peptides as well as the existing amino terminus, employing degenerate polymerase chain reaction primers corresponding to the sequences of the peptides as well as the amino terminus to amplify a fragment from LTA cDNA, and utilizing the resultant probe to obtain full-length enzyme cDNAs from a lambda Zap Express LTA cDNA library. Human 3-OST cDNAs were isolated by searching the expressed sequence tag data bank with the mouse sequence, identifying a partial-length human cDNA and utilizing the clone as a probe to isolate a full-length enzyme cDNA from a lambda TriplEx human brain cDNA library. The expression of wild-type mouse 3-OST as well as protein A-tagged mouse enzyme by transient transfection of COS-7 cells and the expression of both wild-type mouse and human 3-OST by in vitro transcription/translation demonstrate that the two cDNAs directly encode both HSact conversion and 3-OST activities. The mouse 3-OST cDNAs exhibit three different size classes because of a 5'-untranslated region of variable length, which results from the insertion of 0-1629 base pairs (bp) between residues 216 and 217; however, all cDNAs contain the same open reading frame of 933 bp. The length of the 3'-untranslated region ranges from 301 to 430 bp. The nucleic acid sequence of mouse and human 3-OST cDNAs are approximately 85% similar, encoding novel 311- and 307-amino acid proteins of 35,876 and 35,750 daltons, respectively, that are 93% similar. The encoded enzymes are predicted to be intraluminal Golgi residents, presumably interacting via their C-terminal regions with an integral membrane protein. Both 3-OST species exhibit five potential N-glycosylation sites, which account for the apparent discrepancy between the molecular masses of the encoded enzyme (approximately 34 kDa) and the previously purified enzyme (approximately 46 kDa). The two 3-OST species also exhibit approximately 50% similarity with all previously identified forms of the heparan biosynthetic enzyme N-deacetylase/N-sulfotransferase, which suggests that heparan biosynthetic enzymes share a common sulfotransferase domain.

Carotenoids are known to play an important role in health and disease state of living human tissue based on their antioxidant and optical filtering functions. In this study, we show that carotenoids exist in human bone and surrounding fatty tissue both in significant and individually variable concentrations. Measurements of biopsied tissue samples with molecule-specific Raman spectroscopy and high-performance liquid chromatography reveal that all carotenoids that are known to exist in human skin are also present in human bone. This includes all car- otenes, lycopene, β-cryptoxanthin, lutein, and zeaxanthin. We propose quantitative reflection imaging as a non- contact optical method suitable for the measurement of composite carotenoid levels in bone and surrounding tissue exposed during open surgeries such as total knee arthroplasty, and as a proof of concept, demonstrate carotenoid measurements in biopsied bone samples. This will allow one to establish potential correlations between internal tissue carotenoid levels and levels in skin and to potentially use already existing optical skin carotenoid tests as surrogate marker for bone carotenoid status. © 2013 Society of Photo-Optical Instrumentation Engineers (SPIE) (DOI: 10.1117/1.JBO.18.11 .117006)

Led by the federal government, the payers of health care are enacting policies designed to base provider reimbursement on the quality of care they render. This study evaluated and compared patient experiences and satisfaction with nasal decolonization with either nasal povidone-iodine (PI) or nasal mupirocin ointment (MO). A total of 1903 patients were randomized to undergo preoperative nasal decolonization with either nasal MO or PI solution. All randomized patients were also given 2% chlorhexidine gluconate topical wipes. Patients were interviewed prior to discharge to assess adverse events and patient experience with their assigned preoperative antiseptic protocol. Of the 1903 randomized patients, 1679 (88.1%) were interviewed prior to discharge. Of patients receiving PI, 3.4% reported an unpleasant or very unpleasant experience, compared with 38.8% of those using nasal MO ( P P >.05). Being recruited as an active participant in surgical site infection prevention was a positive experience for 87.2% of MO patients and 86.3% of PI patients ( P= .652). Those assigned to receive PI solution preoperatively reported significantly fewer adverse events than the nasal MO group ( P

The study of Fe, Zn and Cd stable isotopes (δ(56)Fe, δ(66)Zn and δ(114)Cd) in seawater is a new field, which promises to elucidate the marine cycling of these bioactive trace metals. However, the analytical challenges posed by the low concentration of these metals in seawater has meant that previous studies have typically required large sample volumes, highly limiting data collection in the oceans. Here, we present the first simultaneous method for the determination of these three isotope systems in seawater, using Nobias PA-1 chelating resin to extract metals from seawater, purification by anion exchange chromatography, and analysis by double spike MC-ICPMS. This method is designed for use on only a single litre of seawater and has blanks of 0.3, 0.06 and

LTA cells synthesize a minor population of heparan sulfate proteoglycans (HSPGact) bearing anticoagulant heparan sulfate (HSact) with a specific monosaccharide sequence that accelerates the action of antithrombin (AT). LTA cells also synthesize a major population of heparan sulfate proteoglycans endowed with nonanticoagulant heparan sulfate (HSinact) lacking the AT-binding site. To investigate the pathway-specific features of HSPGact generation, we established a novel detergent-containing cell-free system with unlabeled and labeled microsomes from wild-type and variant LTA cells, respectively. The unlabeled microsomes provide “HSact conversion activity” that requires 3′-phosphoadenosine 5′-phosphosulfate to convert [35S]HSPGinact into [35S] HSPGact, presumably by sulfation. The reaction kinetics demonstrate that the rate of HSact synthesis is constant over the first 4 h of incubation. During this time, the rate of HSact production is linearly dependent on the amount of unlabeled LTA microsomal protein over a range of 10 to 50 μg as well as on the level of [35S]HS substrate over a range of 0.4 to 4.0 μg, microsomal protein. Compared with labeled microsomes, equivalent or slightly greater levels of HSact were generated from 35S-labeled HSPG, microsomal HS, or cell surface HS, which demonstrates that HSinact is the minimal substrate and that large amounts of HSact precursor exit the Golgi apparatus. Indeed, extensive modification of wild-type LTA cell surface [35S]HS elevated HSact content from 9 to 35%. The hypothesis that microsomal HSact conversion activity predicts the cellular rate of HSact generation was tested with wild-type or variant LTA cells in which production of HSact has been significantly altered by mutagenesis or overexpression of core protein or growth conditions. The data demonstrate that microsomal HSact conversion activity accurately reflects the cellular rate of HSact synthesis over a very wide range of conditions. The possibility that the reduced HSact generation is due to an inhibitor was excluded by mixing experiments. The possibility that reduced HSact generation is caused by decreased levels of HSact precursor was excluded as equivalent levels of HSact were formed from wild-type and variant [35S]HS. Based upon the above data, the LTA cell microsomal HSact conversion activity contains one or more limiting components that kinetically regulate the rate of cellular HSact generation and the levels of HSact precursor in HS greatly exceed HSact production.

An ecto-ATP diphosphohydrolase (ATPDase) was purified to homogeneity from vesiculosomes shed from chicken oviduct. First, the ecto-ATPDase-enriched vesiculosomes were concentrated by filtration, differential centrifugation, and exclusion chromatography. Next, the nonionic detergent, Nonidet P-40, was used to extract the ecto-ATPDase from vesiculosomal membranes, and the solubilized enzyme was further purified by ion exchange (DEAE-Bio-Gel) and lentil-lectin-Sepharose 4B chromatography. In the final stage, immunoaffinity chromatography was utilized to obtain purified ecto-ATPDase. More than 25,000-fold purification was achieved. Specific activity of the purified enzyme was greater than 800 micronol/min/mg of protein with MgATP as the substrate, the highest ever reported for an ATPDase. The enzyme also hydrolyzed other nucleoside triphosphates in the presence of magnesium at similar rates and CaATP and MgADP at lower rates. The molecular mass of the purified glycoprotein was 80 kDa as determined by SDS-polyacrylamide gel electrophoresis and Western blot analysis. Based on its enzymatic properties, the relationship of the chicken oviduct ecto-ATPDase with other reported ATPDases and ecto-ATPases is discussed.

SummaryThe haemostatic system was examined in 2951 men aged 50 to 61 years, clinically free of cardiovascular disease, who were ranked according to a risk score for fatal coronary heart disease (CHD). Risk was judged from their serum cholesterol concentration, systolic blood pressure, body mass index and smoking habit. The status of the factor VH-tissue factor pathway was estimated from the plasma levels of factor VII coagulant activity, factor VII antigen and activated factor VII. Activation of factor IX was assessed from the plasma concentration of factor IX activation peptide. Activity within the common pathway was measured as the plasma concentrations of prothrombin fragment 1+2 and fibrinopeptide A. All 6 markers of haemostatic status were positively and statistically significantly associated with risk, providing further evidence for a hypercoagulable state in men at high risk for fatal CHD. Plasma fibrinogen and serum triglyceride concentrations were also graded positively with risk.