In situ formation of blends by photopolymerization of poly(ethylene glycol) dimethacrylate and polylactide

Blends of cross-linked poly(ethylene glycol) dimethacrylate (PEGDMA) and poly(d,l-lactide) (PLA) were prepared by mixing photoactive PEGDMA (molecular mass: 875 g/mol) and PLA, and subsequently photopolymerizing the mixture with visible light. The effects of PLA molecular mass and mass fraction on the rheological properties of the PEGDMA/PLA mixtures, and on the degree of methacrylate vinyl conversion (DC), as well as blend miscibility, microstructure, mechanical properties, in vitro swelling behavior, and cell responses were studied. PLA-2K (molecular mass: 2096 g/mol) and PLA-63K (molecular mass: 63 000 g/mol) formed miscible and partially miscible blends with cross-linked PEGDMA, respectively. The addition of the PLA-2K did not affect the immediate or post-cure (>24 h) DC of the PEGDMA upon photopolymerization. However, the addition of PLA-63K decreased the immediate DC of the PEGDMA, which can be increased through extending the curing time or post-curing period. Compared to the cross-linked neat PEGDMA and PLA-2K/PEGDMA blends, PLA-63K/PEGDMA blends were significantly stronger, stiffer, and tougher. Both types of blends and the cross-linked PEGDMA swelled when soaked in a phosphate buffered saline (PBS) solution. The attachment and spreading of MCT3-E1 cells increased with increasing PLA-63K content in the blends. The facile and rapid formation of PEGDMA/PLA blends by photopolymerization represents a simple and efficient approach to a class of biomaterials with a broad spectrum of properties.     

Antibody-directed fibrinolysis. An antibody specific for both fibrin and tissue plasminogen activator

An investigation was made to determine whether it is possible to attract tissue plasminogen activator (tPA) to the site of a thrombus by means of an antibody with affinites for both tPA and fibrin. A bispecific antibody conjugate was constructed by cross-linking two monoclonal antibodies: one specific for tPA, the other specific for fibrin. The bispecific antibody enhanced fibrinolysis by capturing tPA at the site of a fibrin deposit. In an in vitro quantitative fibrinolysis assay, the relative fibrinolytic potency of tPA bound to the bispecific antibody was 13 times greater than that of tPA and 200 times greater than that of urokinase. When fibrin was treated with the bispecific antibody before being mixed with tPA, the relative fibrinolytic potency of tPA was enhanced 14-fold. This capture also occurred when the concentration of tPA present in the assay was less than the concentration of tPA present in normal human plasma. In a human plasma clot assay, samples containing both the bispecific antibody and tPA exhibited significantly more lysis than did samples containing tPA alone. In spite of the increased clot lysis effected by the presence of bispecific antibody, there was no significant increase in fibrinogen or alpha 2-antiplasmin degradation at equal tPA concentrations. The ability of the bispecific antibody to concentrate exogenous tPA in vivo was then examined in the rabbit jugular vein model. Systemic infusion of a small amount of tPA (10,000 units) produced no significant increment in thrombolysis over the level of spontaneous lysis (14 +/- 8%). However, the simultaneous infusion of 10,000 units of tPA and 2 mg of bispecific antibody resulted in 42 +/- 14% (p less than 0.01) lysis. These results suggest that a molecule capable of binding both fibrin and tPA with high affinity could enhance thrombolysis in the circulation by capturing endogenous tPA.     

Structural basis for c-di-GMP-mediated inside-out signaling controlling periplasmic proteolysis

The bacterial second messenger bis-(3'-5') cyclic dimeric guanosine monophosphate (c-di-GMP) has emerged as a central regulator for biofilm formation. Increased cellular c-di-GMP levels lead to stable cell attachment, which in Pseudomonas fluorescens requires the transmembrane receptor LapD. LapD exhibits a conserved and widely used modular architecture containing a HAMP domain and degenerate diguanylate cyclase and phosphodiesterase domains. c-di-GMP binding to the LapD degenerate phosphodiesterase domain is communicated via the HAMP relay to the periplasmic domain, triggering sequestration of the protease LapG, thus preventing cleavage of the surface adhesin LapA. Here, we elucidate the molecular mechanism of autoinhibition and activation of LapD based on structure-function analyses and crystal structures of the entire periplasmic domain and the intracellular signaling unit in two different states. In the absence of c-di-GMP, the intracellular module assumes an inactive conformation. Binding of c-di-GMP to the phosphodiesterase domain disrupts the inactive state, permitting the formation of a trans-subunit dimer interface between adjacent phosphodiesterase domains via interactions conserved in c-di-GMP-degrading enzymes. Efficient mechanical coupling of the conformational changes across the membrane is realized through an extensively domain-swapped, unique periplasmic fold. Our structural and functional analyses identified a conserved system for the regulation of periplasmic proteases in a wide variety of bacteria, including many free-living and pathogenic species.     

Seed coat variation in Glycine Willd. subgenus Glycine (Leguminosae) by SEM

Seed of the genusGlycine Willd. typically exhibits a muriculate appearance resulting from adherence to the true seed coat of the perisperm or inner pod wall layer. Thickened cell walls of the perisperm superimpose a reticulate network on the seed coat, the type of network ranging from alveolate to stellate depending on the shape of the perisperm cells. Tubercles distributed at intervals give the seed its roughened appearance. Seed lacking an attached perisperm appears smooth and shiny. Seed morphology of 62 collections representing the six species of the subgenusGlycine is examined in detail to elucidate inter-and intraspecific variability. Seed perisperm pattern appears to be characteristic for each species, but there are exceptions.Glycine canescens F. J. Herrn. andG. clandestina Willd. seeds possess a reticulate network and tubercles of irregular shape, the perisperm appearing granular inG. clandestina. Seeds ofG. latrobeana (Meissn.) Benth. andG. tabacina (Labill.) Benth. lack a distinct network and have stellate tubercles; the perisperm is granular inG. latrobeana and some plants ofG. tabacina. A few collections ofG. clandestina approachG. tabacina in seed appearance.Glycine tomentella Hayata seeds exhibit a regularly alveolate arrangement, while those ofG. falcata Benth. lack a perisperm layer altogether. Variation in seed coat within a species can usually be linked to differences in chromosome number or some aspect of gross morphology. Diploid collections ofG. tomentella (2n = 40) exhibit recognizable differences in seed morphology compared with tetraploids (2n = 80), coincident with other striking dissimilarities in gross morphology. An incompletely attached perisperm is accompanied by aneuploidy in severalG. tomentella accessions, while other 78 and 38 chromosome aneuploids produce normal seeds.     

The contribution of marine snow to the particle food supply of the benthic suspension feeder, Mytilus edulis

In order to examine the importance of the settling of large particles to the food supply and feeding behavior of a benthic culture of the blue mussel, Mytilus edulis, we investigated the tidal dynamics of large (>0.5 mm diameter) marine aggregates, commonly known as marine snow, during three tidal cycles in July 1998 at a shallow, subtidal, low current flow regime site along the coast of Maine (Shorey Cove, Roque Island, Englishman's Bay, Maine).In situ, optically measured marine snow showed a distinct tidal signal displaying an increase in size and abundance through high tide with a peak on the early ebb tide as it settled to the bottom. Marine snow volume ranged an order of magnitude through the tidal cycle, from under 8 to over 80 mm³ l⁻¹. An increase in the in situ marine snow volume corresponded with an increase in benthic mussel feeding activity (from 20% to 60% of maximum exhalant siphon area, which is an estimate of pumping rate) and maximum rates of pseudofeces production by the mussels during periods of low tidal current speeds. In contrast, mussels from the same population feeding on surface waters in shipboard chambers produced no pseudofeces and had high pumping rates (80-100% maximum exhalant siphon area) over the whole tidal period. A second peak in benthic mussel pumping rates also occurred during flood tide.Food quality was lower in the bottom waters due to significantly higher particulate inorganic matter (PIM, >2 mg l⁻¹) when compared with the surface waters. PIM accounted for 95% of the total settled mass flux of 3.4 g m⁻² day⁻¹ measured in sediment traps deployed 1 m off the bottom, with organic carbon representing only 2.5% of the mass flux during the mid-summer conditions. At low-current sites such as Shorey Cove, Roque Island, Maine, the settling of marine snow provides an important additional source of food, albeit of low quality, to benthic populations of blue mussels.     

A cross-continental test of the Enemy Release Hypothesis: leaf herbivory on Acer platanoides (L.) is three times lower in North America than in its native Europe

Acer platanoides (Norway maple) is a widespread native tree species in Europe. It has been introduced to North America where it has often established dense stands in both secondary woodlands and relatively undisturbed mature woodlands. In Europe A. platanoides is also extending its original range, but generally seems to exist at much lower densities. One explanation for the ‘aggressiveness’ of invasive plants such as A. platanoides is that they have left behind pests and diseases which limit their population densities in their native lands (the enemy release hypothesis or ERH). To assess the ERH for Norway maple, a large network of collaborators assessed leaf herbivory rates in populations throughout Europe and North America. We found significantly lower total leaf herbivory (1.6% ± 0.19, n = 21 vs. 7.4% ± 1.94, n = 34) and lower fungal damage (1.0% ± 0.35, n = 13 vs. 3.7% ± 0.85, n = 34) in North America than in Europe over a 2 year period, which is consistent with the predictions of the Enemy Release Hypothesis. Across years, the average total leaf herbivory was significantly correlated with average annual temperature of the site (P < 0.05), although this was mostly due to sites in Europe (P < 0.001), and not sites in North America (P > 0.05). Furthermore, only populations in Europe showed very high levels of herbivory (e.g., nine sites had total leaf herbivory ranging from 10.0 to 51.2% in at least 1 year) or leaf fungal damage (only one site in North America showed high levels of fungal damage in 1 year), suggesting the possibility of more frequent episodic outbreaks in the native range. Leaf herbivory and fungal damage are only two aspects of consumer pressure and we do not know whether the differences reported here are enough to actually elicit release from top-down population control, but such large scale biogeographic differences in herbivory contribute towards understanding exotic invasions. Jonathan M. Adams and Wei Fang—equally contributed as first authors. A list of the participating members of the Transatlantic Acer platanoides Invasion Network is given in the Appendix 3.     

The Irish DAFNE Study Protocol: A cluster randomised trial of group versus individual follow-up after structured education for Type 1 diabetes

Background

Subgroup analyses in randomized trials examine whether effects of interventions differ between subgroups of study populations according to characteristics of patients or interventions. However, findings from subgroup analyses may be misleading, potentially resulting in suboptimal clinical and health decision making. Few studies have investigated the reporting and conduct of subgroup analyses and a number of important questions remain unanswered. The objectives of this study are: 1) to describe the reporting of subgroup analyses and claims of subgroup effects in randomized controlled trials, 2) to assess study characteristics associated with reporting of subgroup analyses and with claims of subgroup effects, and 3) to examine the analysis, and interpretation of subgroup effects for each study's primary outcome.

Methods

We will conduct a systematic review of 464 randomized controlled human trials published in 2007 in the 118 Core Clinical Journals defined by the National Library of Medicine. We will randomly select journal articles, stratified in a 1:1 ratio by higher impact versus lower impact journals. According to 2007 ISI total citations, we consider the New England Journal of Medicine, JAMA, Lancet, Annals of Internal Medicine, and BMJ as higher impact journals. Teams of two reviewers will independently screen full texts of reports for eligibility, and abstract data, using standardized, pilot-tested extraction forms. We will conduct univariable and multivariable logistic regression analyses to examine the association of pre-specified study characteristics with reporting of subgroup analyses and with claims of subgroup effects for the primary and any other outcomes.

Discussion

A clear understanding of subgroup analyses, as currently conducted and reported in published randomized controlled trials, will reveal both strengths and weaknesses of this practice. Our findings will contribute to a set of recommendations to optimize the conduct and reporting of subgroup analyses, and claim and interpretation of subgroup effects in randomized trials.     

Treatment of Cryptococcal Meningitis in KwaZulu-Natal,South Africa

Background

Cryptococcal meningitis (CM) remains a leading cause of death for HIV-infected individuals in sub-Saharan Africa. Improved treatment strategies are needed if individuals are to benefit from the increasing availability of antiretroviral therapy. We investigated the factors associated with mortality in routine care in KwaZulu-Natal, South Africa.

Methodology/Principal Findings

A prospective year long, single-center, consecutive case series of individuals diagnosed with cryptococcal meningitis 190 patients were diagnosed with culture positive cryptococcal meningitis, of whom 186 were included in the study. 52/186 (28.0%) patients died within 14 days of diagnosis and 60/186 (32.3%) had died by day 28. In multivariable cox regression analysis, focal neurology (aHR 11 95%C.I. 3.08–39.3, P<0.001), diastolic blood pressure <60 mmHg (aHR 2.37 95%C.I. 1.11–5.04, P = 0.025), concurrent treatment for tuberculosis (aHR 2.11 95%C.I. 1.02–4.35, P = 0.044) and use of fluconazole monotherapy (aHR 3.69 95% C.I. 1.74–7.85, P<0.001) were associated with increased mortality at 14 and 28 days.

Conclusions

Even in a setting where amphotericin B is available, mortality from cryptococcal meningitis in this setting is high, particularly in the immediate period after diagnosis. This highlights the still unmet need not only for earlier diagnosis of HIV and timely access to treatment of opportunistic infections, but for better treatment strategies of cryptococcal meningitis.     

Biosecurity-based interventions and strategies to reduce Campylobacter spp. on poultry farms

The prevention and control of Campylobacter colonization of poultry flocks are important public health strategies for the control of human campylobacteriosis. A critical review of the literature on interventions to control Campylobacter in poultry on farms was undertaken using a systematic approach. Although the focus of the review was on aspects appropriate to the United Kingdom poultry industry, the research reviewed was gathered from worldwide literature. Multiple electronic databases were employed to search the literature, in any language, from 1980 to September 2008. A primary set of 4,316 references was identified and scanned, using specific agreed-upon criteria, to select relevant references related to biosecurity-based interventions. The final library comprised 173 references. Identification of the sources of Campylobacter in poultry flocks was required to inform the development of targeted interventions to disrupt transmission routes. The approach used generally involved risk factor-based surveys related to culture-positive or -negative flocks, usually combined with a structured questionnaire. In addition, some studies, either in combination or independently, undertook intervention trials. Many of these studies were compromised by poor design, sampling, and statistical analysis. The evidence for each potential source and route of transmission on the poultry farm was reviewed critically, and the options for intervention were considered. The review concluded that, in most instances, biosecurity on conventional broiler farms can be enhanced and this should contribute to the reduction of flock colonization. However, complementary, non-biosecurity-based approaches will also be required in the future to maximize the reduction of Campylobacter-positive flocks at the farm level.     

Longitudinal molecular epidemiological study of thermophilic campylobacters on one conventional broiler chicken farm

Improved understanding of the ecology and epidemiology of Campylobacter in the poultry farm environment is key to developing appropriate farm-based strategies for preventing flock colonization. The sources of Campylobacter causing broiler flock colonization were investigated on one poultry farm and its environment, from which samples were obtained on three occasions during each of 15 crop cycles. The farm was adjacent to a dairy farm, with which there was a shared concreted area and secondary entrance. There was considerable variation in the Campylobacter status of flocks at the various sampling times, at median ages of 20, 26, and 35 days, with 3 of the 15 flocks remaining negative at slaughter. Campylobacters were recoverable from various locations around the farm, even while the flock was Campylobacter negative, but the degree of environmental contamination increased substantially once the flock was positive. Molecular typing showed that strains from house surroundings and the dairy farm were similar to those subsequently detected in the flock and that several strains intermittently persisted through multiple crop cycles. The longitudinal nature of the study suggested that bovine fecal Campylobacter strains, initially recovered from the dairy yard, may subsequently colonize poultry. One such strain, despite being repeatedly recovered from the dairy areas, failed to colonize the concomitant flock during later crop cycles. The possibility of host adaptation of this strain was investigated with 16-day-old chickens experimentally exposed to this strain naturally present in, or spiked into, bovine feces. Although the birds became colonized by this infection model, the strain may preferentially infect cattle. The presence of Campylobacter genotypes in the external environment of the poultry farm, prior to their detection in broiler chickens, confirms the horizontal transmission of these bacteria into the flock and highlights the risk from multispecies farms.