Apatite-Polymer Composite Particles for Controlled Delivery of BMP-2: In Vitro Release and Cellular Response

Bone morphogenetic protein-2 (BMP-2) has the ability to induce osteoblast differentiation of undifferentiated cells, resulting in the healing of skeletal defects when delivered with a suitable carrier. We have applied a versatile delivery platform comprising a novel composite of two biomaterials with proven track records – apatite and poly(lactic-co-glycolic acid) (PLGA) – to the delivery of BMP-2. Sustained release of this growth factor was tuned with variables that affect polymer degradation and/or apatite dissolution, such as polymer molecular weight, polymer composition, apatite loading, and apatite particle size. The effect of released BMP-2 on C3H10T1/2 murine pluripotent mesenchymal cells was assessed by tracking the expression of osteoblastic makers, alkaline phosphatase (ALP) and osteocalcin. Release media collected over 100 days induced elevated ALP activity in C3H10T1/2 cells. The expression of osteocalcin was also upregulated significantly. These results demonstrated the potential of apatite-PLGA composite particles for releasing protein in bioactive form over extended periods of time.

Primary succession in man-made wetlands: biodiversity, structure and dynamics of macrofaunal assemblages

Man-made wetlands are often created to compensate for the loss or degradation of natural wetlands, but little is known about the processes taking place in these artificial environments, especially at the community level. Throughout this thesis, we have assessed the phenomena of primary succession over different time (short-, mid- and long-term) and spatial scales (local, regional, interregional levels), applying different approaches (taxonomic and functional) and subject groups (invertebrates and amphibians). Our main findings regarding time scales show a 3-phase successional pattern in Mediterranean man-made wetlands’ communities, where at the short term (1 year) colonization processes dominate; at mid term perspectives (2 to 7 years) succession signs begin to be conspicuous, and later on (≥ 10 years) parameters such as species richness reach an asymptote. At that moment, some biological strategies dominate, and biodiversity surrogates indicate that communities are indistinct between man-made and natural wetlands. Regarding spatial effects, we corroborated that both local and regional factors affect the establishing communities. Particularly, the low hydrological stability of the Mediterranean region has enhanced biological traits favoring resilience and resistance to disturbances when comparing Mediterranean and cold temperate aquatic communities. Even within the Mediterranean region, low levels of hydrological stability have significant effects on the successional dynamics. In these cases, local communities are highly nested within regional natural ones, and so are not able to make net contributions to regional richness. We also showed the influence of the regional pool of recruiters over local communities, both in the case of invertebrates and amphibians. Especially for the latter group, man-made Mediterranean temporary ponds (MTPs) can play an important role in their conservation.

Variation of the mixing state of Saharan dust particles with atmospheric transport

Mineral dust is an important aerosol species in the Earth’s atmosphere and has a major source within North Africa, of which the Sahara forms the major part. Aerosol Time of Flight Mass Spectrometry (ATOFMS) is first used to determine the mixing state of dust particles collected from the land surface in the Saharan region, showing low abundance of species such as nitrate and sulphate internally mixed with the dust mineral matrix. These data are then compared with the ATOFMS single particle mass spectra of Saharan dust particles detected in the marine atmosphere in the vicinity of the Cape Verde islands, which are further compared with those from particles with longer atmospheric residence sampled at a coastal station at Mace Head, Ireland. Saharan dust particles collected near the Cape Verde Islands showed increased internally mixed nitrate but no sulphate, whilst Saharan dust particles collected on the coast of Ireland showed a very high degree of internally mixed secondary species including nitrate, sulphate and methanesulphonate. This uptake of secondary species will change the pH and hygroscopic properties of the aerosol dust and thus can influence the budgets of other reactive gases, as well as influencing the radiative properties of the particles and the availability of metals for dissolution.

The role of weeds in supporting biological diversity within crop fields

Weeds are major constraints on crop production, yet as part of the primary producers within farming systems, they may be important components of the agroecosystem. Using published literature, the role of weeds in arable systems for other above-ground trophic levels are examined. In the UK, there is evidence that weed flora have changed over the past century, with some species declining in abundance, whereas others have increased. There is also some evidence for a decline in the size of arable weed seedbanks. Some of these changes reflect improved agricultural efficiency, changes to more winter-sown crops in arable rotations and the use of more broad-spectrum herbicide combinations. Interrogation of a database of records of phytophagous insects associated with plant species in the UK reveals that many arable weed species support a high diversity of insect species. Reductions in abundances of host plants may affect associated insects and other taxa. A number of insect groups and farmland birds have shown marked population declines over the past 30 years. Correlational studies indicate that many of these declines are associated with changes in agricultural practices. Certainly reductions in food availability in winter and for nestling birds in spring are implicated in the declines of several bird species, notably the grey partridge, Perdix perdix . Thus weeds have a role within agroecosystems in supporting biodiversity more generally. An understanding of weed competitivity and the importance of weeds for insects and birds may allow the identification of the most important weed species. This may form the first step in balancing the needs for weed control with the requirements for biodiversity and more sustainable production methods.

Oxidation of biogenic and water-soluble compounds in aqueous and organic aerosol droplets by ozone: a kinetic and product analysis approach using laser Raman tweezers

The results of an experimental study into the oxidative degradation of proxies for atmospheric aerosol are presented. We demonstrate that the laser Raman tweezers method can be used successfully to obtain uptake coeffcients for gaseous oxidants on individual aqueous and organic droplets, whilst the size and composition of the droplets is simultaneously followed. A laser tweezers system was used to trap individual droplets containing an unsaturated organic compound in either an aqueous or organic ( alkane) solvent. The droplet was exposed to gas- phase ozone and the reaction kinetics and products followed using Raman spectroscopy. The reactions of three different organic compounds with ozone were studied: fumarate anions, benzoate anions and alpha pinene. The fumarate and benzoate anions in aqueous solution were used to represent components of humic- like substances, HULIS; a alpha- pinene in an alkane solvent was studied as a proxy for biogenic aerosol. The kinetic analysis shows that for these systems the diffusive transport and mass accommodation of ozone is relatively fast, and that liquid- phase di. ffusion and reaction are the rate determining steps. Uptake coe. ffcients, g, were found to be ( 1.1 +/- 0.7) x 10(-5), ( 1.5 +/- 0.7) x 10 (-5) and ( 3.0 - 7.5) x 10 (-3) for the reactions of ozone with the fumarate, benzoate and a- pinene containing droplets, respectively. Liquid- phase bimolecular rate coe. cients for reactions of dissolved ozone molecules with fumarate, benzoate and a- pinene were also obtained: k(fumarate) = ( 2.7 +/- 2) x 10 (5), k(benzoate) = ( 3.5 +/- 3) x 10 (5) and k(alpha-pinene) = ( 1-3) x 10(7) dm(3) mol (-1) s (- 1). The droplet size was found to remain stable over the course of the oxidation process for the HULIS- proxies and for the oxidation of a- pinene in pentadecane. The study of the alpha- pinene/ ozone system is the first using organic seed particles to show that the hygroscopicity of the particle does not increase dramatically over the course of the oxidation. No products were detected by Raman spectroscopy for the reaction of benzoate ions with ozone. One product peak, consistent with aqueous carbonate anions, was observed when following the oxidation of fumarate ions by ozone. Product peaks observed in the reaction of ozone with alpha- pinene suggest the formation of new species containing carbonyl groups.

Effect of oxidized low-density lipoprotein on differential gene expression in primary human endothelial cells

Oxidative modification of low-density lipoprotein (LDL) plays an important role in the initiation and progression of atherosclerosis. It has been proposed that the biological action of oxidized LDL (ox-LDL) may be partially attributed to its effect on a shift of the pattern of gene expression in endothelial cells. To examine the transcriptional response to ox-LDL, we applied cDNA array technology to cultured primary human endothelial cells challenged with oxidized human LDL. A twofold or greater difference in the expression of a particular gene was considered a significant difference in transcript abundance. Seventy-eight of the 588 genes analyzed were differentially expressed in response to the treatment. Ox-LDL significantly affected the expression of genes encoding for transcription factors, cell receptors, growth factors, adhesion molecules, extracellular matrix proteins, and enzymes involved in cholesterol metabolism. The alteration of the expression pattern of several genes was substantiated post hoc using RT-PCR. The experimental strategy identified several novel ox-LDL-sensitive genes associated with a "response to injury" providing a conceptual background to be utilized for future studies addressing the molecular basis of the early stages of atherogenesis.

Genistein affects the expression of genes involved in blood pressure regulation and angiogenesis in primary human endothelial cells

Background: Several lines of evidence suggest that the dietary isoflavone genistein (Gen) has beneficial effects with regard to cardiovascular disease and in particular on aspects related to blood pressure and angiogenesis. The biological action of Gen may be, at Least in part, attributed to its ability to affect cell signalling and response. However, so far, most of the molecular mechanisms underlying the activity of Gen in the endothelium are unknown. Methods and results: To examine the transcriptional response to 2.5 mu M Gen on primary human endothelial cells (HUVEC), we applied cDNA array technology both under baseline condition and after treatment with the pro-atherogenic stimulus, copper-oxidized LDL. The alteration of the expression patterns of individual transcripts was substantiated using either RT-PCR or Northern blotting. Gen significantly affected the expression of genes encoding for proteins centrally involved in the vascular tone such as endothelin-converting enzyme-1, endothetin-2, estrogen related receptor a and atria[ natriuretic peptide receptor A precursor. Furthermore, Gen countered the effect of oxLDL on mRNA levels encoding for vascular endothelial growth factor receptor 165, types 1 and 2. Conclusions: Our data indicate that physiologically achievable levels of Gen change the expression of mRNA encoding for proteins involved in the control of blood pressure under baseline conditions and reduce the angiogenic response to oxLDL in the endothelium. (c) 2005 Elsevier B.V. All rights reserved.

Numerical modeling of inhaled charged aerosol deposition in human airways

A new numerical modeling of inhaled charge aerosol has been developed based on a modified Weibel's model. Both the velocity profiles (slug and parabolic flows) and the particle distributions (uniform and parabolic distributions) have been considered. Inhaled particles are modeled as a dilute dispersed phase flow in which the particle motion is controlled by fluid force and external forces acting on particles. This numerical study extends the previous numerical studies by considering both space- and image-charge forces. Because of the complex computation of interacting forces due to space-charge effect, the particle-mesh (PM) method is selected to calculate these forces. In the PM technique, the charges of all particles are assigned to the space-charge field mesh, for calculating charge density. The Poisson's equation of the electrostatic potential is then solved, and the electrostatic force acting on individual particle is interpolated. It is assumed that there is no effect of humidity on charged particles. The results show that many significant factors also affect the deposition, such as the volume of particle cloud, the velocity profile and the particle distribution. This study allows a better understanding of electrostatic mechanism of aerosol transport and deposition in human airways.

Ice formation and development in aged, wintertime cumulus over the UK: observations and modelling

In situ high resolution aircraft measurements of cloud microphysical properties were made in coordination with ground based remote sensing observations of a line of small cumulus clouds, using Radar and Lidar, as part of the Aerosol Properties, PRocesses And InfluenceS on the Earth's climate (APPRAISE) project. A narrow but extensive line (~100 km long) of shallow convective clouds over the southern UK was studied. Cloud top temperatures were observed to be higher than −8 °C, but the clouds were seen to consist of supercooled droplets and varying concentrations of ice particles. No ice particles were observed to be falling into the cloud tops from above. Current parameterisations of ice nuclei (IN) numbers predict too few particles will be active as ice nuclei to account for ice particle concentrations at the observed, near cloud top, temperatures (−7.5 °C). The role of mineral dust particles, consistent with concentrations observed near the surface, acting as high temperature IN is considered important in this case. It was found that very high concentrations of ice particles (up to 100 L−1) could be produced by secondary ice particle production providing the observed small amount of primary ice (about 0.01 L−1) was present to initiate it. This emphasises the need to understand primary ice formation in slightly supercooled clouds. It is shown using simple calculations that the Hallett-Mossop process (HM) is the likely source of the secondary ice. Model simulations of the case study were performed with the Aerosol Cloud and Precipitation Interactions Model (ACPIM). These parcel model investigations confirmed the HM process to be a very important mechanism for producing the observed high ice concentrations. A key step in generating the high concentrations was the process of collision and coalescence of rain drops, which once formed fell rapidly through the cloud, collecting ice particles which caused them to freeze and form instant large riming particles. The broadening of the droplet size-distribution by collision-coalescence was, therefore, a vital step in this process as this was required to generate the large number of ice crystals observed in the time available. Simulations were also performed with the WRF (Weather, Research and Forecasting) model. The results showed that while HM does act to increase the mass and number concentration of ice particles in these model simulations it was not found to be critical for the formation of precipitation. However, the WRF simulations produced a cloud top that was too cold and this, combined with the assumption of continual replenishing of ice nuclei removed by ice crystal formation, resulted in too many ice crystals forming by primary nucleation compared to the observations and parcel modelling.

Sensitivity analysis of an ocean carbon cycle model in the North Atlantic: an investigation of parameters affecting the air-sea CO2 flux, primary production and export of detritus

The sensitivity of the biological parameters in a nutrient-phytoplankton-zooplankton-detritus (NPZD) model in the calculation of the air-sea CO2 flux, primary production and detrital export is analysed. We explore the effect on these outputs of variation in the values of the twenty parameters that control ocean ecosystem growth in a 1-D formulation of the UK Met Office HadOCC NPZD model used in GCMs. We use and compare the results from one-at-a-time and all-at-a-time perturbations performed at three sites in the EuroSITES European Ocean Observatory Network: the Central Irminger Sea (60° N 40° W), the Porcupine Abyssal Plain (49° N 16° W) and the European Station for Time series in the Ocean Canary Islands (29° N 15° W). Reasonable changes to the values of key parameters are shown to have a large effect on the calculation of the air-sea CO2 flux, primary production, and export of biological detritus to the deep ocean. Changes in the values of key parameters have a greater effect in more productive regions than in less productive areas. The most sensitive parameters are generally found to be those controlling well-established ocean ecosystem parameterisations widely used in many NPZD-type models. The air-sea CO2 flux is most influenced by variation in the parameters that control phytoplankton growth, detrital sinking and carbonate production by phytoplankton (the rain ratio). Primary production is most sensitive to the parameters that define the shape of the photosynthesis-irradiance curve. Export production is most sensitive to the parameters that control the rate of detrital sinking and the remineralisation of detritus.

Modeling ocean primary production: sensitivity to spectral resolution of attenuation and absorption of light

Modeling the vertical penetration of photosynthetically active radiation (PAR) through the ocean, and its utilization by phytoplankton, is fundamental to simulating marine primary production. The variation of attenuation and absorption of light with wavelength suggests that photosynthesis should be modeled at high spectral resolution, but this is computationally expensive. To model primary production in global 3d models, a balance between computer time and accuracy is necessary. We investigate the effects of varying the spectral resolution of the underwater light field and the photosynthetic efficiency of phytoplankton (α∗), on primary production using a 1d coupled ecosystem ocean turbulence model. The model is applied at three sites in the Atlantic Ocean (CIS (∼60°N), PAP (∼50°N) and ESTOC (∼30°N)) to include the effect of different meteorological forcing and parameter sets. We also investigate three different methods for modeling α∗ – as a fixed constant, varying with both wavelength and chlorophyll concentration [Bricaud, A., Morel, A., Babin, M., Allali, K., Claustre, H., 1998. Variations of light absorption by suspended particles with chlorophyll a concentration in oceanic (case 1) waters. Analysis and implications for bio-optical models. J. Geophys. Res. 103, 31033–31044], and using a non-spectral parameterization [Anderson, T.R., 1993. A spectrally averaged model of light penetration and photosynthesis. Limnol. Oceanogr. 38, 1403–1419]. After selecting the appropriate ecosystem parameters for each of the three sites we vary the spectral resolution of light and α∗ from 1 to 61 wavebands and study the results in conjunction with the three different α∗ estimation methods. The results show modeled estimates of ocean primary productivity are highly sensitive to the degree of spectral resolution and α∗. For accurate simulations of primary production and chlorophyll distribution we recommend a spectral resolution of at least six wavebands if α∗ is a function of wavelength and chlorophyll, and three wavebands if α∗ is a fixed value.

The role of sky conditions on gross primary production in a mixed deciduous forest

The role of different sky conditions on diffuse PAR fraction (ϕ), air temperature (Ta), vapor pressure deficit (vpd) and GPP in a deciduous forest is investigated using eddy covariance observations of CO2 fluxes and radiometer and ceilometer observations of sky and PAR conditions on hourly and growing season timescales. Maximum GPP response occurred under moderate to high PAR and ϕ and low vpd. Light response models using a rectangular hyperbola showed a positive linear relation between ϕ and effective quantum efficiency (α = 0.023ϕ + 0.012, r2 = 0.994). Since PAR and ϕ are negatively correlated, there is a tradeoff between the greater use efficiency of diffuse light and lower vpd and the associated decrease in total PAR available for photosynthesis. To a lesser extent, light response was also modified by vpd and Ta. The net effect of these and their relation with sky conditions helped enhance light response under sky conditions that produced higher ϕ. Six sky conditions were classified from cloud frequency and ϕ data: optically thick clouds, optically thin clouds, mixed sky (partial clouds within hour), high, medium and low optical aerosol. The frequency and light responses of each sky condition for the growing season were used to predict the role of changing sky conditions on annual GPP. The net effect of increasing frequency of thick clouds is to decrease GPP, changing low aerosol conditions has negligible effect. Increases in the other sky conditions all lead to gains in GPP. Sky conditions that enhance intermediate levels of ϕ, such as thin or scattered clouds or higher aerosol concentrations from volcanic eruptions or anthropogenic emissions, will have a positive outcome on annual GPP, while an increase in cloud cover will have a negative impact. Due to the ϕ/PAR tradeoff and since GPP response to changes in individual sky conditions differ in sign and magnitude, the net response of ecosystem GPP to future sky conditions is non-linear and tends toward moderation of change.

Estimating the direct aerosol radiative perturbation: impact of ocean surface representation and aerosol non-sphericity

Atmospheric aerosols are now actively studied, in particular because of their radiative and climate impacts. Estimations of the direct aerosol radiative perturbation, caused by extinction of incident solar radiation, usually rely on radiative transfer codes and involve simplifying hypotheses. This paper addresses two approximations which are widely used for the sake of simplicity and limiting the computational cost of the calculations. Firstly, it is shown that using a Lambertian albedo instead of the more rigorous bidirectional reflectance distribution function (BRDF) to model the ocean surface radiative properties leads to large relative errors in the instantaneous aerosol radiative perturbation. When averaging over the day, these errors cancel out to acceptable levels of less than 3% (except in the northern hemisphere winter). The other scope of this study is to address aerosol non-sphericity effects. Comparing an experimental phase function with an equivalent Mie-calculated phase function, we found acceptable relative errors if the aerosol radiative perturbation calculated for a given optical thickness is daily averaged. However, retrieval of the optical thickness of non-spherical aerosols assuming spherical particles can lead to significant errors. This is due to significant differences between the spherical and non-spherical phase functions. Discrepancies in aerosol radiative perturbation between the spherical and non-spherical cases are sometimes reduced and sometimes enhanced if the aerosol optical thickness for the spherical case is adjusted to fit the simulated radiance of the non-spherical case.

Impact of the modal aerosol scheme GLOMAP-mode on aerosol forcing in the Hadley Centre Global Environmental Model

The Hadley Centre Global Environmental Model (HadGEM) includes two aerosol schemes: the Coupled Large-scale Aerosol Simulator for Studies in Climate (CLASSIC), and the new Global Model of Aerosol Processes (GLOMAP-mode). GLOMAP-mode is a modal aerosol microphysics scheme that simulates not only aerosol mass but also aerosol number, represents internally-mixed particles, and includes aerosol microphysical processes such as nucleation. In this study, both schemes provide hindcast simulations of natural and anthropogenic aerosol species for the period 2000–2006. HadGEM simulations of the aerosol optical depth using GLOMAP-mode compare better than CLASSIC against a data-assimilated aerosol re-analysis and aerosol ground-based observations. Because of differences in wet deposition rates, GLOMAP-mode sulphate aerosol residence time is two days longer than CLASSIC sulphate aerosols, whereas black carbon residence time is much shorter. As a result, CLASSIC underestimates aerosol optical depths in continental regions of the Northern Hemisphere and likely overestimates absorption in remote regions. Aerosol direct and first indirect radiative forcings are computed from simulations of aerosols with emissions for the year 1850 and 2000. In 1850, GLOMAP-mode predicts lower aerosol optical depths and higher cloud droplet number concentrations than CLASSIC. Consequently, simulated clouds are much less susceptible to natural and anthropogenic aerosol changes when the microphysical scheme is used. In particular, the response of cloud condensation nuclei to an increase in dimethyl sulphide emissions becomes a factor of four smaller. The combined effect of different 1850 baselines, residence times, and abilities to affect cloud droplet number, leads to substantial differences in the aerosol forcings simulated by the two schemes. GLOMAP-mode finds a presentday direct aerosol forcing of −0.49Wm−2 on a global average, 72% stronger than the corresponding forcing from CLASSIC. This difference is compensated by changes in first indirect aerosol forcing: the forcing of −1.17Wm−2 obtained with GLOMAP-mode is 20% weaker than with CLASSIC. Results suggest that mass-based schemes such as CLASSIC lack the necessary sophistication to provide realistic input to aerosol-cloud interaction schemes. Furthermore, the importance of the 1850 baseline highlights how model skill in predicting present-day aerosol does not guarantee reliable forcing estimates. Those findings suggest that the more complex representation of aerosol processes in microphysical schemes improves the fidelity of simulated aerosol forcings.

Natural aerosol direct and indirect radiative effects

Natural aerosol plays a significant role in the Earth’s system due to its ability to alter the radiative balance of the Earth. Here we use a global aerosol microphysics model together with a radiative transfer model to estimate radiative effects for five natural aerosol sources in the present-day atmosphere: dimethyl sulfide (DMS), sea-salt, volcanoes, monoterpenes, and wildfires. We calculate large annual global mean aerosol direct and cloud albedo effects especially for DMS-derived sulfate (–0.23 Wm–2 and –0.76 Wm–2, respectively), volcanic sulfate (–0.21 Wm–2 and –0.61 Wm–2) and sea-salt (–0.44 Wm–2 and –0.04 Wm–2). The cloud albedo effect responds nonlinearly to changes in emission source strengths. The natural sources have both markedly different radiative efficiencies and indirect/direct radiative effect ratios. Aerosol sources that contribute a large number of small particles (DMS-derived and volcanic sulfate) are highly effective at influencing cloud albedo per unit of aerosol mass burden.