Employing citizen science to enhance active and healthy ageing in urban environments.

Engaging older residents in problem definition and solution-building is key to the success of place-based initiatives endeavouring to increase the age-friendliness of urban environments. This study employed the Our Voice framework, engaging older adult citizen scientists (n = 14) and community stakeholders (n = 15) across the city of Birmingham, UK. With the aim of identifying urban features impacting age friendliness and co-producing recommendations for improving local urban areas, citizen scientists participated in 12 technology-enabled walkability assessments, three in-person discussion groups, two one-to-one online discussions, and two workshops with community stakeholders. Together, citizen scientists co-produced 12 local and six city-wide recommendations. These recommendations were embedded into an implementation framework based on workshop discussions to identify age-friendly pathways in urban environments.

A novel cell line panel reveals non-genetic mediators of platinum resistance and phenotypic diversity in high grade serous ovarian cancer.

OBJECTIVES: Resistance to cancer therapy is an enduring challenge and accurate and reliable preclinical models are lacking. We interrogated this unmet need using high grade serous ovarian cancer (HGSC) as a disease model. METHODS: We created five in vitro and two in vivo platinum-resistant HGSC models and characterised the entire cell panel via whole genome sequencing, RNASeq and creation of intraperitoneal models. RESULTS: Mutational signature analysis indicated that platinum-resistant cell lines evolved from a pre-existing ancestral clone but a unifying mutational cause for drug resistance was not identified. However, cisplatin-resistant and carboplatin-resistant cells evolved recurrent changes in gene expression that significantly overlapped with independent samples obtained from multiple patients with relapsed HGSC. Gene Ontology Biological Pathways (GOBP) related to the tumour microenvironment, particularly the extracellular matrix, were repeatedly enriched in cisplatin-resistant cells, carboplatin-resistant cells and also in human resistant/refractory samples. The majority of significantly over-represented GOBP however, evolved uniquely in either cisplatin- or carboplatin-resistant cell lines resulting in diverse intraperitoneal behaviours that reflect different clinical manifestations of relapsed human HGSC. CONCLUSIONS: Our clinically relevant and usable models reveal a key role for non-genetic factors in the evolution of chemotherapy resistance. Biological pathways relevant to the extracellular matrix were repeatedly expressed by resistant cancer cells in multiple settings. This suggests that recurrent gene expression changes provide a fitness advantage during platinum therapy and also that cancer cell-intrinsic mechanisms influence the tumour microenvironment during the evolution of drug resistance. Candidate genes and pathways identified here could reveal therapeutic opportunities in platinum-resistant HGSC.

The Role of Urban Environments in Promoting Active and Healthy Aging: A Systematic Scoping Review of Citizen Science Approaches.

Promoting active and healthy aging in urban spaces requires environments with diverse, age-friendly characteristics. This scoping review investigated the associations between urban characteristics and active and healthy aging as identified by citizen science (CS) and other participatory approaches. Using a systematic scoping review procedure, 23 articles employing a CS or participatory approach (participant age range: 54-98 years) were reviewed. An inductive and deductive thematic analysis was completed to (a) identify local urban barriers and facilitators and (b) map them against the World Health Organization (WHO) Checklist of Essential Features of Age-Friendly Cities. A new Citizen Science Appraisal Tool (CSAT) was developed to evaluate the quality of CS and other participatory approaches included in the reviewed articles. A range of interconnected urban barriers and facilitators was generated by residents across the personal (e.g. perceived safety), environmental (e.g. unmaintained infrastructure), socio-cultural (e.g. cross-cultural activities), economic (e.g. affordable housing) and political (e.g. governmental support to migrant communities) domains. Mapping the barriers and facilitators to the WHO age-friendly checklist underscored the checklist's relevance and elucidated the need to explore barriers for migrant and cross-cultural communities and neighborhood development and alterations. The CSAT demonstrated strengths related to active engagement of residents and study outcomes leading to real-world implications. To advance the potential of CS to enrich our understanding of age-friendly environments, employing co-production to enhance relevance and sustainability of outcomes is an important strategy. Overall, employing CS highlighted the value of systematically capturing the experiences of older adults within studies aimed at promoting active and healthy aging.

Cleavage of nicotinamide adenine dinucleotide by the ribosome-inactivating protein from Momordica charantia.

The interaction of momordin, a type 1 ribosome-inactivating protein from Momordica charantia, with NADP(+) and NADPH has been investigated by X-ray diffraction analysis of complexes generated by co-crystallization and crystal soaking. It is known that the proteins of this family readily cleave the adenine-ribose bond of adenosine and related nucleotides in the crystal, leaving the product, adenine, bound to the enzyme active site. Surprisingly, the nicotinamide-ribose bond of oxidized NADP(+) is cleaved, leaving nicotinamide bound in the active site in the same position but in a slightly different orientation to that of the five-membered ring of adenine. No binding or cleavage of NADPH was observed at pH 7.4 in these experiments. These observations are in accord with current views of the enzyme mechanism and may contribute to ongoing searches for effective inhibitors.

Immunocytochemical analysis of synaptic proteins provides new insights into diabetes-mediated plasticity in the rat hippocampus.

The hippocampus, an important integration center for learning and memory in the mammalian brain, undergoes neurological changes in response to a variety of stimuli that are suggestive of ongoing synaptic reorganization. Accordingly, the aim of this study was to identify markers of synaptic plasticity using rapid and reliable techniques such as radioimmunocytochemistry and confocal microscopy, thereby providing a "birds-eye view" of the whole hippocampus under hypercorticosteronemic conditions. The regulation of microtubule-associated protein 2, synaptophysin and postsynaptic density-95 was examined in two different animal models of hypercorticosteronemia: corticosterone administration and streptozotocin-induced diabetes using both a short-term (1 week) and long-term (5 weeks) treatment. Glucocorticoids and/or hyperglycemia increased synaptophysin expression in CA1, CA3 and the dentate gyrus, regions that exhibit synaptic plasticity in response to glucocorticoid exposure. In these models, postsynaptic density-95 expression increased in the CA3 region, particularly in the diabetic rats, while microtubule-associated protein 2 exhibited more selective changes. Fluoro-Jade histochemistry did not detect neuronal damage, suggesting that glucocorticoids and/or hyperglycemia induce plastic and not irreversible neuronal changes at these time points. Collectively, these results demonstrate that changes in the expression and distribution of synaptic proteins provide another measure of synaptic plasticity in the rat hippocampus in response to glucocorticoid exposure, changes that may accompany or contribute to neuroanatomical, neurochemical, and behavioral changes observed in experimental models of type 1 diabetes.

The role of the hippocampus in trace conditioning: temporal discontinuity or task difficulty?

It is well established that the hippocampal formation is critically involved in the acquisition of trace memories, a paradigm in which the conditioned (CS) and unconditioned stimuli (US) are separated by a temporal gap (Solomon et al., 1986). The structure is reportedly not critical for the acquisition of delay memories, where the CS and the US overlap in time (Berger & Orr, 1983; Schmaltz & Theios, 1972). Based on these results, it is often stated that the hippocampus is involved in "filling the gap" or otherwise associating the two stimuli in time. However, in addition to the presence of a temporal gap, there are other differences between trace and delay conditioning. The most apparent difference is that animals require many more trials to learn the trace task, and thus it is inherently more difficult than the delay task. Here, we tested whether the hippocampus was critically involved in delay conditioning, if it was rendered more difficult such that the rate of acquisition was shifted to be analogous to trace conditioning. Groups of rats received excitotoxic lesions to the hippocampus, sham lesions or were left intact. Using the same interstimulus intervals (ISI), control animals required more trials to acquire the trace than the delay task. As predicted, animals with hippocampal lesions were impaired during trace conditioning but not delay conditioning. However, when the delay task was rendered more difficult by extending the ISI (a long delay task), animals with hippocampal lesions were impaired. In addition, once the lesioned animal learned the association between the CS and the US during delay conditioning, it could learn and perform the trace CR. Thus, the role of the hippocampus in classical conditioning is not limited to learning about discontiguous events in time and space; rather the structure can become engaged simply as a function of task difficulty.

The genome of the natural genetic engineer Agrobacterium tumefaciens C58.

The 5.67-megabase genome of the plant pathogen Agrobacterium tumefaciens C58 consists of a circular chromosome, a linear chromosome, and two plasmids. Extensive orthology and nucleotide colinearity between the genomes of A. tumefaciens and the plant symbiont Sinorhizobium meliloti suggest a recent evolutionary divergence. Their similarities include metabolic, transport, and regulatory systems that promote survival in the highly competitive rhizosphere; differences are apparent in their genome structure and virulence gene complement. Availability of the A. tumefaciens sequence will facilitate investigations into the molecular basis of pathogenesis and the evolutionary divergence of pathogenic and symbiotic lifestyles.

Haemophilus ducreyi inhibits phagocytosis by U-937 cells, a human macrophage-like cell line.

Haemophilus ducreyi is a gram-negative obligate human pathogen that causes the genital ulcer disease chancroid. Chancroid lesions are deep necrotic ulcers with an immune cell infiltrate that includes macrophages. Despite the presence of these phagocytic cells, chancroid ulcers can persist for months and live H. ducreyi can be isolated from these lesions. To analyze the interaction of H. ducreyi with macrophages, we investigated the ability of H. ducreyi strain 35000 to adhere to, invade, and survive within U-937 cells, a human macrophage-like cell line. We found that although H. ducreyi strain 35000 adhered efficiently to U-937 cells, few bacteria were internalized, suggesting that H. ducreyi avoids phagocytosis by human macrophages. The few bacteria that were phagocytosed in these experiments were rapidly killed. We also found that H. ducreyi inhibits the phagocytosis of a secondary target (opsonized sheep red blood cells). Antiphagocytic activity was found in logarithmic, stationary-phase, and plate-grown cultures and was associated with whole, live bacteria but not with heat-killed cultures, sonicates, or culture supernatants. Phagocytosis was significantly inhibited after a 15-min exposure to H. ducreyi, and a multiplicity of infection of approximately 1 CFU per macrophage was sufficient to cause a significant reduction in phagocytosis by U-937 cells. Finally, all of nine H. ducreyi strains tested were antiphagocytic, suggesting that this is a common virulence mechanism for this organism. This finding suggests a mechanism by which H. ducreyi avoids killing and clearance by macrophages in chancroid lesions and inguinal lymph nodes.

The contribution of adrenal and reproductive hormones to the opposing effects of stress on trace conditioning in males versus females.

Exposure to an acute stressful experience facilitates classical conditioning in male rats but impairs conditioning in female rats (T. J. Shors, C. Lewczyk, M. Paczynski, P. R. Mathew, & J. Pickett, 1998; G. E. Wood & T. J. Shors, 1998). The authors report that these effects extend to performance on the hippocampal-dependent task of trace conditioning. The stress-induced impairment of conditioning in females was evident immediately, 24 hr and 48 hr after stress, depending on the stage of estrus. Moreover, the effect could be reactivated days later by reexposure to the stressful context. Corticosterone levels correlated with overall performance in males but not in females. Unlike the effect seen in males, adrenalectomy did not prevent the stress-induced effect on conditioning in females. These data indicate that exposure to the same experience can have opposite effects on learning in males versus females and that these opposing effects are mediated by differing hormonal systems.

The Bvg accessory factor (Baf) enhances pertussis toxin expression in Escherichia coli and is essential for Bordetella pertussis viability.

Pertussis toxin expression in the Gram-negative respiratory pathogen, Bordetella pertussis, is regulated by the BvgAS two-component system. Previous studies suggested that an additional gene encoding a Bvg accessory factor (Baf) was required, along with BvgAS, for expression of a ptx-lacZ fusion in Escherichia coli grown in rich medium. However, other studies showed that BvgAS is sufficient for ptx-lacZ expression in minimal medium. Here we show that Baf acts with BvgAS to further increase ptx-lacZ expression in E. coli grown in minimal media and this is concomitant with a two-fold increase in BvgA protein levels. Gene replacement experiments show that baf is essential for viability of B. pertussis, suggesting that Baf affects the expression of other genes in addition to ptx.

The modulation of Pavlovian memory.

Exposure to stressful experiences as well as sex differences in the brain are known to influence the acquisition of new memories. This review focuses on acquisition of two types of Pavlovian learning paradigms: hippocampal-independent delay conditioning and hippocampal-dependent trace conditioning and their modulation by exposure to stressful experience and sex differences in the brain. We concentrate on two sets of findings: the first is that exposure to an acute stressful experience enhances Pavlovian conditioning in the male rat, while exposure to the very same experience dramatically impairs conditioning in female rat. The sexually-opposed effects of stress on conditioning are mediated by differing hormonal substrates (adrenal versus ovarian steroids) and possibly by differing anatomical and biochemical pathways. The second set of findings is that training with hippocampal-dependent trace conditioning enhances the survival of newly generated neurons in the adult hippocampal formation. The same amount of training with hippocampal-independent delay conditioning does not affect their survival. In addition, females acquire the trace task faster than males and generate more new neurons. As with the stress effects on learning, these sex effects are influenced by hormonal status. It is our contention that identifying the hormonal and neuronal processes that modulate associative memory formation will provide insight into the processes of memory formation itself.

Target cell range of Haemophilus ducreyi hemolysin and its involvement in invasion of human epithelial cells.

Haemophilus ducreyi, the causative agent of chancroid, produces a hemolysin, whose role in virulence is not well defined. To assess the possible role of hemolysin in pathogenesis, we evaluated its target cell range by using wild-type H. ducreyi 35000, nonhemolytic mutants with the hemolysin structural gene deleted, and isogenic strains expressing different amounts of hemolytic activity. The cytotoxicity of the various cell types was assessed by quantitating the release of lactate dehydrogenase into culture supernatants as a measure of cell lysis. In these experiments, human foreskin fibroblasts, human foreskin epithelial cells, and, to a lesser extent, HEp-2 cells were lysed by H. ducreyi hemolysin. Hemolysin also lysed human blood mononuclear cells and immune system cell lines including U937 macrophage-like cells, T lymphocytes, and B lymphocytes. In contrast, human polymorphonuclear leukocytes were not sensitive to hemolysin under the conditions tested. We also analyzed the effect of hemolysin on invasion of human epithelial cells and found that H. ducreyi strains expressing cloned hemolysin genes showed a 10-fold increase in invasion compared to the control strain. These data support the hypothesis that the H. ducreyi hemolysin is important in the pathogenesis of chancroid and may contribute to ulcer formation, invasion of epithelial cells, and evasion of the immune response.

Prevalence of, antibody response to, and immunity induced by Haemophilus ducreyi hemolysin.

Haemophilus ducreyi, the etiologic agent of chancroid, a genital ulcer disease, produces a cell-associated hemolysin whose role in virulence is not well defined. Hemolysin is encoded by two genes, hhdA and hhdB, which, based on their homology to Serratia marcescens shlA and shlB genes, are believed to encode the hemolysin structural protein and a protein required for secretion and modification of this protein, respectively. In this study, we determined the prevalence and expression of the hemolysin genes in 90 H. ducreyi isolates obtained from diverse geographic locations from 1952 to 1996 and found that all strains contained DNA homologous to the hhdB and hhdA genes. In addition, all strains expressed a hemolytic activity. We also determined that hemolysin is expressed in vivo and is immunogenic, as indicated by the induction of antibodies to hemolysin in both the primate and rabbit disease models as well as in human patients with naturally acquired chancroid. Wild-type strain 35000 and isogenic hemolysin-negative mutants showed no difference in lesion development in the temperature-dependent rabbit model. However, immunization of rabbits with the purified hemolysin protein reduced the recovery of wild-type H. ducreyi, but not hemolysin-negative mutants, from lesions. Our study indicates that hemolysin is a possible candidate for vaccine development due to its immunogenicity, expression in vitro and in vivo by most, if not all, strains, and the effect of immunization on reducing the recovery of viable H. ducreyi in experimental disease in rabbits.

Determination of deoxynivalenol in white flour, whole wheat flour, and bran by solid-phase extraction/liquid chromatography: interlaboratory study.

A liquid chromatographic (LC) method for determining deoxynivalenol (DON) in white flour, whole wheat flour, and bran at or above the U.S. Food and Drug Administration advisory level of 1 microgram/g was evaluated by an interlaboratory study. Test samples of processed wheat (flour and bran) were extracted by blending with acetonitrile-water (84 + 16). Extracts were filtered and passed through a solid-phase extraction (SPE) column. The eluate was then chromatographed on a reversed-phase LC column with a water-methanol gradient. DON was measured at 220 nm. Naturally contaminated white flour, whole wheat flour, and bran samples and spiking solutions of DON to be added to the 3 commodities at 0.5, 1.0, and 2.0 micrograms/g were sent to 4 collaborators in Kansas, Louisiana, Missouri, and Washington states. Three collaborators completed the study. Average recoveries of DON from the 3 commodities spiked at 0.5, 1.0, and 2.0 micrograms/g were 94, 87, and 97%, respectively. Within-laboratory relative standard deviations for repeatability (RSDr) ranged from 3.1 to 21.7% and between-laboratory relative standard deviations for reproducibility (RSDR) ranged from 10.8 to 38.7%. On the basis of the results of this study, the SPE/LC method for DON in white flour, whole wheat flour, and bran was adopted as a peer-verified method by AOAC INTERNATIONAL.

Stress facilitates classical conditioning in males, but impairs classical conditioning in females through activational effects of ovarian hormones.

Exposure to restraint and brief intermittent tailshocks facilitates associative learning of the classical conditioned eyeblink response in male rats. Based on evidence of sex differences in learning and responses to stressful events, we investigated sexually dimorphic effects of a stressor of restraint and intermittent tailshock on classical eyeblink conditioning 24 h after stressor cessation. Our results indicate that exposure to the acute stressor had diametrically opposed effects on the rate of acquisition of the conditioned response in male vs. female rats. Exposure to the stressor facilitated acquisition of the conditioned response in males, whereas exposure to the same stressful event dramatically impaired acquisition in females. We further demonstrate that the stress-induced impairment in female conditioning is dependent on the presence of ovarian hormones. Conditioning of stressed sham-ovariectomized females was significantly impaired relative to the unstressed controls, whereas conditioning in stressed ovariectomized females was not impaired. We present additional evidence that estrogen mediates the stress-induced impairment in female acquisition. Females administered sesame oil vehicle and then stressed were significantly impaired relative to their unstressed controls, whereas females administered the estrogen antagonist tamoxifen prior to stress were not impaired. In summary, these results indicate that exposure to the same aversive event can induce opposite behavioral responses in males vs. females. These effects underscore sex differences in associative learning and emotional responding, and implicate estrogen in the underlying neuronal mechanism.

Identification of Btr-regulated genes using a titration assay. Search for a role for this transcriptional regulator in the growth and virulence of Bordetella pertussis.

Bordetella pertussis is the causative agent of the respiratory disease pertussis or whopoping cough. Btr, an oxygen-responsive transcriptional regulator of B. pertussis, is homologous to the FNR protein of E. coli. Using a murine respiratory model, we observed in the present study that Btr is important in growth and survival of B. pertussis in vivo. A titration assay was developed that identified genes containing Btr binding sites including B. pertussis sodB and btr, E. coli aspA and a new B. pertussis gene, brg1. The brg1 gene encodes a protein similar to the LysR family of transcriptional regulators and its expression is activated threefold by Btr under anaerobic growth conditions but unaffected by Btr aerobically. The nucleotide sequence flanking brg1 encodes proteins with similarity to various metabolic enzymes. Putative overlapping promoters and a Btr binding site (FNR box) were identified in the DNA sequence between brg1 and the adjacent genes. These intervening sequences may represent sites for regulation by Btr and Brg1.

Determination and survey of deoxynivalenol in white flour, whole wheat flour, and bran.

A liquid chromatographic (LC) method for determining deoxynivalenol (DON) in white flour, whole wheat flour, and bran was developed. A 25 g test portion was extracted with acetonitrile-water (84 + 16), and the extract was filtered and applied to a column containing a combination of charcoal, Celite, and other adsorbents. The eluate was then chromatographed on a silica-based, reversed-phase LC column by using a gradient of water and methanol. DON was measured at 220 nm. Average recoveries of DON from white flour, whole wheat flour, and bran spiked at 1 microgram/g were 88, 86, and 85%, respectively. The limit of determination of the method was < 0.5 micrograms/g. A total of 562 wheat-based products from the 1993 crop year were collected by 21 U.S. Food and Drug Administration District Offices and analyzed by this method in Kansas City, Seattle, and New Orleans District Laboratories. The numbers of samples with DON contamination > or = 1 microgram/g from 163 bran, 272 white flour, 90 whole wheat flour, and 37 miscellaneous test samples were 20, 28, 14, and 2, respectively. About 52, 50, 40, and 27% of the same test samples were contaminated with DON at levels > 0.01 micrograms/g.

Contribution of stress and gender to exploratory preferences for familiar versus unfamiliar conspecifics.

An apparatus for measuring the exploratory preferences of rats for familiar and unfamiliar conspecifics in a novel environment was designed. The exploratory behavior of males and females was compared and contrasted to that elicited in response to an acute aversive event. Sprague-Dawley male and female rats were exposed to restraint and 60, 1 s, 1 mA tailshocks and returned to their home cage. Either 2 or 24 h later, they were placed in a novel environment with a familiar cage-mate and an unfamiliar conspecific of the same sex. Relative to unstressed controls and females, males stressed 2 h previously decreased the exploration of the unfamiliar conspecific, exhibiting a rapid decrease over the course of the trial. In response to the stressor, however both sexes, however, decreased the exploration of the familiar conspecific, decreased their overall activity, and returned preferentially to their starting quadrant. None of these stress-induced effects were evident 24 h later upon the first or second exposure to the apparatus. Thus, exposure to the stressor transiently increased perseveration and decreased activity in males and females, but only decreased the exploration of novel conspecifics in males. These results indicate that a number of behavioral responses to stressors are conserved across gender, but those relating to novelty are more pronounced in males.

Identification of a Bordetella pertussis regulatory factor required for transcription of the pertussis toxin operon in Escherichia coli.

Transcription of the pertussis toxin operon (ptx) is positively regulated in Bordetella pertussis by the bvgAS locus. However, a ptx-lacZ transcriptional fusion in Escherichia coli cannot be activated by bvgAS in trans. This suggests that an additional factor(s) is required for transcription of ptx. A gene encoding a Bvg accessory factor (Baf) was identified by its ability to activate an E. coli ptx-lacZ fusion in the presence of bvgAS. The expression of ptx-lacZ was decreased by the addition of 40 mM MgSO4, a compound that also modulates ptx expression in B. pertussis. Baf alone did not activate expression of an E. coli fhaB-lacZ fusion, nor did it increase expression of fhaB-lacZ in trans with bvgAS. The gene encoding Baf was localized, sequenced, and found to produce a novel 28-kDa protein. Sequences homologous to B. pertussis baf were identified in Bordetella bronchiseptica and Bordetella parapertussis but not in Bordetella avium. When an additional copy of baf was integrated into the chromosome of BC75, a B. pertussis mutant that produces a low level of pertussis toxin, pertussis toxin production was partially complemented in the cointegrate strain.