Restoring Carboxylates on Highly Modified Alginates Improves Gelation, Tissue Retention and Systemic Capture.

Alginate hydrogels are gaining traction for use in drug delivery, regenerative medicine, and as tissue engineered scaffolds due to their physiological gelation conditions, high tissue biocompatibility, and wide chemical versatility. Traditionally, alginate is decorated at the carboxyl group to carry drug payloads, peptides, or proteins. While low degrees of substitution do not cause noticeable mechanical changes, high degrees of substitution can cause significant losses to alginate properties including complete loss of calcium cross-linking. While most modifications used to decorate alginate deplete the carboxyl groups, we propose that alginate modifications that replenish the carboxyl groups could overcome the loss in gel integrity and mechanics. In this report, we demonstrate that restoring carboxyl groups during functionalization maintains calcium cross-links as well as hydrogel shear-thinning and self-healing properties. In addition, we demonstrate that alginate hydrogels modified to a high degree with azide modifications that restore the carboxyl groups have improved tissue retention at intramuscular injection sites and capture blood-circulating cyclooctynes better than alginate hydrogels modified with azide modifications that deplete the carboxyl groups. Taken together, alginate modifications that restore carboxyl groups could significantly improve alginate hydrogel mechanics for clinical applications. STATEMENT OF SIGNIFICANCE: Chemical modification of hydrogels provides a powerful tool to regulate cellular adhesion, immune response, and biocompatibility with local tissues. Alginate, due to its biocompatibility and easy chemical modification, is being explored for tissue engineering and drug delivery. Unfortunately, modifying alginate to a high degree of substitution consumes carboxyl group, which are necessary for ionic gelation, leading to poor hydrogel crosslinking. We introduce alginate modifications that restore the alginate's carboxyl groups. We demonstrate that modifications that reintroduce carboxyl groups restore gelation and improve gel mechanics and tissue retention. In addition to contributing to a basic science understanding of hydrogel properties, we anticipate our approach will be useful to create tissue engineered scaffolds and drug delivery platforms.

The signature of competition in ecomorphological traits across the avian radiation.

Competition for shared resources represents a fundamental driver of biological diversity. However, the tempo and mode of phenotypic evolution in deep-time has been predominantly investigated using trait evolutionary models which assume that lineages evolve independently from each other. Consequently, the role of species interactions in driving macroevolutionary dynamics remains poorly understood. Here, we quantify the prevalence for signatures of competition between related species in the evolution of ecomorphological traits across the bird radiation. We find that mechanistic trait models accounting for the effect of species interactions on phenotypic divergence provide the best fit for the data on at least one trait axis in 27 out of 59 clades ranging between 21 and 195 species. Where it occurs, the signature of competition generally coincides with positive species diversity-dependence, driven by the accumulation of lineages with similar ecologies, and we find scarce evidence for trait-dependent or negative diversity-dependent phenotypic evolution. Overall, our results suggest that the footprint of interspecific competition is often eroded in long-term patterns of phenotypic diversification, and that other selection pressures may predominantly shape ecomorphological diversity among extant species at macroevolutionary scales.

An outbreak of meticillin-resistant Staphylococcus aureus colonization in a neonatal intensive care unit: use of a case-control study to investigate and control it and lessons learnt.

AIM: To describe the investigation and management of a meticillin-resistant Staphylococcus aureus (MRSA) outbreak on a neonatal intensive care unit (NICU) and the lessons learnt. METHODS: This was an outbreak report and case-control study conducted in a 40-cot NICU in a tertiary referral hospital and included all infants colonized/infected with gentamicin-resistant MRSA. INTERVENTION: Standard infection-control measures including segregation of infants, barrier precautions, enhanced cleaning, assessment of staff practice including hand hygiene, and increased MRSA screening of infants were implemented. Continued MRSA acquisitions led to screening of all NICU staff. A case-control study was performed to assess staff contact with colonized babies and inform the management of the outbreak. FINDINGS: Eight infants were colonized with MRSA (spa type t2068), one of whom subsequently developed an MRSA bacteraemia. MRSA colonization was significantly associated with lower gestational age; lower birthweight and with being a twin. Three nurses were MRSA colonized but only one nurse (45) was colonized with MRSA spa type t2068. Multivariable logistic regression analysis identified being cared for by nurse 45 as an independent risk factor for MRSA colonization. CONCLUSIONS: Lack of accurate recording of which nurses looked after which infants (and when) made identification of the risk posed by being cared for by particular nurses difficult. If this had been clearer, it may have enabled earlier identification of the colonized nurse, avoiding subsequent cases. This study highlights the benefit of using a case-control study, which showed that most nurses had no association with colonized infants.

Comparative efficacy of antimicrobial treatments in dairy cows at dry-off to prevent new intramammary infections during the dry period or clinical mastitis during early lactation: a systematic review and network meta-analysis.

A systematic review and network meta-analysis were conducted to assess the relative efficacy of antimicrobial therapy given to dairy cows at dry-off. Eligible studies were controlled trials assessing the use of antimicrobials compared to no treatment or an alternative treatment, and assessed one or more of the following outcomes: incidence of intramammary infection (IMI) at calving, incidence of IMI during the first 30 days in milk (DIM), or incidence of clinical mastitis during the first 30 DIM. Databases and conference proceedings were searched for relevant articles. The potential for bias was assessed using the Cochrane Risk of Bias 2.0 algorithm. From 3480 initially identified records, 45 trials had data extracted for one or more outcomes. Network meta-analysis was conducted for IMI at calving. The use of cephalosporins, cloxacillin, or penicillin with aminoglycoside significantly reduced the risk of new IMI at calving compared to non-treated controls (cephalosporins, RR = 0.37, 95% CI 0.23-0.65; cloxacillin, RR = 0.55, 95% CI 0.38-0.79; penicillin with aminoglycoside, RR = 0.42, 95% CI 0.26-0.72). Synthesis revealed challenges with a comparability of outcomes, replication of interventions, definitions of outcomes, and quality of reporting. The use of reporting guidelines, replication among interventions, and standardization of outcome definitions would increase the utility of primary research in this area.

Comparative efficacy of teat sealants given prepartum for prevention of intramammary infections and clinical mastitis: a systematic review and network meta-analysis.

A systematic review and network meta-analysis were conducted to assess the relative efficacy of internal or external teat sealants given at dry-off in dairy cattle. Controlled trials were eligible if they assessed the use of internal or external teat sealants, with or without concurrent antimicrobial therapy, compared to no treatment or an alternative treatment, and measured one or more of the following outcomes: incidence of intramammary infection (IMI) at calving, IMI during the first 30 days in milk (DIM), or clinical mastitis during the first 30 DIM. Risk of bias was based on the Cochrane Risk of Bias 2.0 tool with modified signaling questions. From 2280 initially identified records, 32 trials had data extracted for one or more outcomes. Network meta-analysis was conducted for IMI at calving. Use of an internal teat sealant (bismuth subnitrate) significantly reduced the risk of new IMI at calving compared to non-treated controls (RR = 0.36, 95% CI 0.25-0.72). For comparisons between antimicrobial and teat sealant groups, concerns regarding precision were seen. Synthesis of the primary research identified important challenges related to the comparability of outcomes, replication and connection of interventions, and quality of reporting of study conduct.

Correlates of rate heterogeneity in avian ecomorphological traits.

Heterogeneity in rates of trait evolution is widespread, but it remains unclear which processes drive fast and slow character divergence across global radiations. Here, we test multiple hypotheses for explaining rate variation in an ecomorphological trait (beak shape) across a globally distributed group (birds). We find low support that variation in evolutionary rates of species is correlated with life history, environmental mutagenic factors, range size, number of competitors, or living on islands. Indeed, after controlling for the negative effect of species' age, 80% of variation in species-specific evolutionary rates remains unexplained. At the clade level, high evolutionary rates are associated with unusual phenotypes or high species richness. Taken together, these results imply that macroevolutionary rates of ecomorphological traits are governed by both ecological opportunity in distinct adaptive zones and niche differentiation among closely related species.

Contribution of parenting to complex syntax development in preschool children with developmental delays or typical development.

BACKGROUND: Despite studies of how parent-child interactions relate to early child language development, few have examined the continued contribution of parenting to more complex language skills through the preschool years. The current study explored how positive and negative parenting behaviours relate to growth in complex syntax learning from child age 3 to age 4 years, for children with typical development or developmental delays (DDs). METHODS: Participants were children with or without DD (N = 60) participating in a longitudinal study of development. Parent-child interactions were transcribed and coded for parenting domains and child language. Multiple regression analyses were used to identify the contribution of parenting to complex syntax growth in children with typical development or DD. RESULTS: Analyses supported a final model, F(9,50) = 11.90, P < .001, including a significant three-way interaction between positive parenting behaviours, negative parenting behaviours and child delay status. This model explained 68.16% of the variance in children's complex syntax at age 4. Simple two-way interactions indicated differing effects of parenting variables for children with or without DD. CONCLUSIONS: Results have implications for understanding of complex syntax acquisition in young children, as well as implications for interventions.

The effect of carbon dioxide flow rate on the euthanasia of laboratory mice.

Laboratory rodents are commonly euthanized by exposure to gradually increasing concentrations of carbon dioxide (CO2). Current recommended flow rates range between 10 and 30% chamber vol/min and result in insensibility before exposure to painful concentrations (<40%). However, this method causes dyspnea, indicated by deep, rapid breathing. In humans dyspnea is associated with a negative affective experience. Sensations of dyspnea may explain why rodents find CO2 concentrations >3% aversive. This study aimed to assess the effect of CO2 flow rates on time between the onset of dyspnea and various measures of insensibility (recumbency, loss of the righting reflex and loss of the pedal withdrawal reflex) to identify flow rates that minimize the potential experience of dyspnea. The results of this study indicate that a flow rate of 50% chamber vol/min, while holding the CO2 cage concentration just below 40%, minimizes the interval between the onset of labored breathing and recumbency. Using a 50% flow rate this interval averaged (± SE) 30.3 ± 2.9 s versus 49.7 ± 2.9 s at 20% chamber vol/min (F3,22 = 7.83, P = 0.0013). Similarly, the interval between the onset of labored breathing and loss of the righting reflex averaged 38.2 ± 2.4 s at a flow rate of 50% versus 59.2 ± 2.4 s at 20% chamber vol/min of CO2 (F3,22 = 13.62, P < 0.0001). We conclude that higher flow rates reduce the duration of dyspnea, but even at the highest flow rate mice experience more than 30 s between the onset of dyspnea and the most conservative estimate of insensibility.

Patent ductus arteriosus evaluation by serial echocardiography in preterm infants.

BACKGROUND: Patent ductus arteriosus (PDA) is associated with morbidity and mortality in premature neonates. METHODS: The effect of serial echocardiography performed by a neonatologist and early targeted medical PDA treatment was evaluated and compared to historical controls. One hundred ninety-two infants <1,500 g were included and 45 infants had a PDA. RESULTS: Serial echocardiography allowed significantly earlier identification and treatment of PDA versus awaiting the evolution of clinical signs. Severe intraventricular haemorrhage and ventilator days were significantly decreased in the studied cohort following the introduction of echocardiography. In addition, hospital stay was also reduced in the non-PDA group and other outcomes were unchanged. CONCLUSION: Serial echocardiography for PDA evaluation, performed by a neonatologist trained in echo, may reduce morbidity in preterm infant.

Biogeological Raman spectroscopic studies of Antarctic lacustrine sediments.

Analysis of lacustrine sediments is an accepted method for deciphering the palaeoenvironment of a lake's catchment area, as each strata of the sediment gives information about the rock type it was eroded from and also the state of the lake, i.e. oxic or anoxic. Antarctica has long been accepted as a putative analogue for Mars, so the analysis of Antarctic material may give results that can be compared to sediments on Mars. Raman spectroscopy has been selected as the method of analysis as it does not destroy the sample, can be used in situ and requires very little sample preparation. It is a suitable method for analysing both inorganic and organic matter and a miniature spectrometer is currently being developed for use in the field. The results from the spectrometers can serve as a guide for analysing sediments on Mars. It has been shown that Raman spectroscopy can detect and differentiate between oxic and anoxic sediments. Both 1064 and 785 nm wavelengths are suitable for laser excitation of organic and inorganic matter.

Gliotoxin is a dual inhibitor of farnesyltransferase and geranylgeranyltransferase I with antitumor activity against breast cancer in vivo.

Gliotoxin is a natural mycotoxin with immunosuppressive and antimicrobial activity. Inhibition of farnesyltransferase (IC50 80 microM) and geranylgeranyltransferase I (IC50 17 microM) stimulated interest in the potential antitumor activity of this epidithiodioxopiperazine. Gliotoxin inhibited proliferation of six breast cancer cell lines in culture with mean +/- SD IC50 289 +/- 328 microM (range 38-985 microM); intracellular farnesylation of Lamin B and geranylgeranylation of Rap1A were inhibited in a dose-dependent manner. In randomized controlled studies using the N-methyl-N-nitrosourea rat mammary carcinoma model, gliotoxin had pronounced antitumor activity in vitro and little systemic toxicity when administered to 10 animals at 10 mg/kg by subcutaneous injection weekly for 4 wk compared with 10 controls. Single doses up to 25 mg/kg were well tolerated. The present studies confirm that gliotoxin is a dual inhibitor of farnesyltransferase and geranylgeranyltransferase I with pronounced antitumor activity and favorable toxicity profile against breast cancer in vitro and in vivo.

Characterization of a mechanism-based inhibitor of NAD(P)H:quinone oxidoreductase 1 by biochemical, X-ray crystallographic, and mass spectrometric approaches.

We report the characterization of 5-methoxy-1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione (ES936) as a mechanism-based inhibitor of NQO1. Inactivation of NQO1 by ES936 was time- and concentration-dependent and required the presence of a pyridine nucleotide cofactor consistent with a need for metabolic activation. That ES936 was an efficient inhibitor was demonstrated in these studies by the low partition ratio (1.40 +/- 0.03). The orientation of ES936 in the active site of NQO1 was examined by X-ray crystallography and found to be opposite to that observed for other indolequinones acting as substrates. ES936 was oriented in such a manner that, after enzymatic reduction and loss of a nitrophenol leaving group, a reactive iminium species was located in close proximity to nucleophilic His 162 and Tyr 127 and Tyr 129 residues in the active site. To determine if ES936 was covalently modifying NQO1, ES936-treated protein was analyzed by electrospray ionization liquid chromatography/mass spectrometry (ESI-LC/MS). The control NQO1 protein had a mass of 30864 +/- 6 Da (n = 20, theoretical, 30868.6 Da) which increased by 217 Da after ES936 treatment (31081 +/- 7 Da, n = 20) in the presence of NADH. The shift in mass was consistent with adduction of NQO1 by the reactive iminium derived from ES936 (M + 218 Da). Chymotryptic digestion of the protein followed by LC/MS analysis located a tetrapeptide spanning amino acids 126-129 which was adducted with the reactive iminium species derived from ES936. LC/MS/MS analysis of the peptide fragment confirmed adduction of either Tyr 127 or Tyr 129 residues. This work demonstrates that ES936 is a potent mechanism-based inhibitor of NQO1 and may be a useful tool in defining the role of NQO1 in cellular systems and in vivo.

Site-specific regulation of oestrogen receptor-alpha and -beta by oestradiol in human adipose tissue.

AIM: To examine the expression of oestrogen receptors alpha and beta (ERalpha and ERbeta) and their regulation by 17beta-oestradiol (E2) in stromal cells and adipocytes from human subcutaneous (s.c.) and omental (o.m.) adipose tissue. METHODS: Subcutaneous and o.m. abdominal adipose tissues were obtained from 10 women (mean age 63.5 +/- 4.8 years; mean weight 75.6 +/- 6.7 kg) undergoing elective or cosmetic surgery. Immunohistochemistry and RT-PCR analysis were used to detect the presence of ERalpha and ERbeta. The regulation of ERalpha and ERbeta by E(2) (10(-7) M to 10(-9) M) was examined using Western immunoblotting analysis in both s.c. and o.m. stromal cells and mature adipocytes cultured in serum-free, phenol red-free medium. RESULTS: Immunostaining of s.c. and o.m. adipose tissue showed that the ER subtypes were localized predominantly within the nucleus. Western analysis demonstrated that E2 treatments differentially altered ERalpha and ERbeta expression in s.c. and o.m. adipocytes. In s.c. and o.m. stromal cells, E(2) (10(-8) M) produced a significant up regulation relative to control of 66 kDa ERalpha (s.c.:1.87 +/- 0.22; o.m.:1.97 +/- 0.17; p < 0.05) and 60 kDa ERbeta (s.c.:1.66 +/- 0.3; o.m.: 1.68 +/- 0.16; p < 0.05). In s.c. adipocytes, however, ERalpha expression significantly decreased with E(2) 10(-8) M relative to control while ERbeta expression increased (ERalpha 0.58 +/- 0.06, ERbeta: 1.47 +/- 0.11; p < 0.05). In o.m. adipocytes, the inhibition of ERalpha with E(2) was not observed (ERalpha 1.86 +/- 0.36, ERbeta:1.03 +/- 0.15, p < 0.05) CONCLUSIONS: ERalpha and ERbeta are expressed but differentially regulated by E(2) in s.c. and o.m. adipocytes and stromal cells. The upregulation of ERbeta by E(2) suggests that E(2) maintains the expression of these receptors. The feed-back inhibition of ERalpha expression by E(2) in s.c. but not o.m. adipocytes observed in vitro is consistent with the data from ERalpha knock out mice where s.c. fat is increased. Selective ER modulators may have different effects in different adipose sites.

Structure-based development of anticancer drugs: complexes of NAD(P)H:quinone oxidoreductase 1 with chemotherapeutic quinones.

BACKGROUND: NAD(P)H:quinone acceptor oxidoreductase (QR1) protects animal cells from the deleterious and carcinogenic effects of quinones and other electrophiles. Remarkably, the same enzyme activates cancer prodrugs that become cytotoxic only after two-electron reduction. QR1's ability to bioactivate quinones and its elevated expression in many human solid tumors makes this protein an excellent target for enzyme-directed drug development. Until now, structural analysis of the mode of binding of chemotherapeutic compounds to QR1 was based on model building using the structures of complexes with simple substrates; no structure of complexes of QR1 with chemotherapeutic prodrugs had been reported. RESULTS: Here we report the high-resolution crystal structures of complexes of QR1 with three chemotherapeutic prodrugs: RH1, a water-soluble homolog of dimethylaziridinylbenzoquinone; EO9, an aziridinylindolequinone; and ARH019, another aziridinylindolequinone. The structures, determined to resolutions of 2.0 A, 2.5 A, and 1.86 A, respectively, were refined to R values below 21% with excellent geometry. CONCLUSIONS: The structures show that compounds can bind to QR1 in more than one orientation. Surprisingly, the two aziridinylindolequinones bind to the enzyme in different orientations. The results presented here reveal two new factors that must be taken into account in the design of prodrugs targeted for activation by QR1: the enzyme binding site is highly plastic and changes to accommodate binding of different substrates, and homologous drugs with different substituents may bind to QR1 in different orientations. These structural insights provide important clues for the optimization of chemotherapeutic compounds that utilize this reductive bioactivation pathway.

Indolequinone antitumor agents: correlation between quinone structure and rate of metabolism by recombinant human NAD(P)H:quinone oxidoreductase. Part 2.

A series of indolequinones bearing various functional groups has been synthesized, and the effects of substituents on the metabolism of the quinones by recombinant human NAD(P)H:quinone oxidoreductase (NQO1) were studied. Indolequinones were selected for study on the basis of the X-ray crystal structure of the human enzyme, and were designed to probe the effect of substituents particularly at N-1. Metabolism of the quinones by NQO1 revealed that, in general, compounds with electron-withdrawing groups at the indole 3-position were among the best substrates, and that groups larger than methyl at N-1 are clearly tolerated. Compounds with a leaving group at the 3-indolyl methyl position generally inactivated the enzyme. The toxicity toward human colon carcinoma cells with either no detectable activity (BE-WT) or high NQO1 activity (BE-NQ) was also studied in representative quinones. The most toxic compounds were those with a leaving group at the C-3 position; these compounds were 1.1-5.3-fold more toxic to the BE-NQ than the BE-WT cells.

The development of primary health care in Jamaica

The development of primary health care in Jamaica is explored, tracing its early roots to the abolition of the slave trade, the collapse of estate-based services after emancipation and the subsequent establishment of the Island Medical Services in 1875. Most development in the health sector occurred after World War I in response to the high infectious disease mortality rates. The Rockefeller Foundation was asked to assist with the control of hookworm, tuberculosis, malaria and yaws. Its recommendations led to the growth of public health programmes (e.g. environmental health, public health nursing, community midwifery) alongside community-based curative services run by hospitals. The most significant period of development occurred in the 1970s when the various vertical programmes were integrated into the current primary care system. Jamaica was integral in the development of the World Health Organization's Alma Ata Declaration on Primary Health Care, tabling the "Jamaican Perspective on Primary Health Care" which set out its goal that all citizens should be within 10 miles walking distance of a primary health care facility. At the close of the twentieth century, the health reform process led to the development of regional health authorities aimed at integrating the management of primary and secondary care under four Regional Boards of Health. This has led to a change in the role of the central Ministry of Health to one of policy-making, health promotion, setting standards, monitoring and evaluation of the quality of health care.

The development of primary health care in Jamaica

The development of primary health care in Jamaica is explored, tracing its early roots to the abolition of the slave trade, the collapse of estate-based services after emancipation and the subsequent establishment of the Island Medical Services in 1875. Most development in the health sector occurred after World War I in response to the high infectious disease mortality rates. The Rockefeller Foundation was asked to assist with the control of hookworm, tuberculosis, malaria and yaws. Its recommendations led to the growth of public health programmes (e.g. environmental health, public health nursing, community midwifery) alongside community-based curative services run by hospitals. The most significant period of development occurred in the 1970s when the various vertical programmes were integrated into the current primary care system. Jamaica was integral in the development of the World Health Organization's Alma Ata Declaration on Primary Health Care, tabling the "Jamaican Perspective on Primary Health Care" which set out its goal that all citizens should be within 10 miles walking distance of a primary health care facility. At the close of the twentieth century, the health reform process led to the development of regional health authorities aimed at integrating the management of primary and secondary care under four Regional Boards of Health. This has led to a change in the role of the central Ministry of Health to one of policy-making, health promotion, setting standards, monitoring and evaluation of the quality of health care.(AU)