Measurement of carotenoid levels in human serum and a catalog of the lutein conformation populations from semi-empirical calculations

Lutein is a principal constituent of the human macular pigment. This study is composed of two projects. The first studies the conformational geometries of lutein and its potential adaptability in biological systems. The second is a study of the response of human subjects to lutein supplements. Using semi-empirical parametric method 3 (PM3) and density functional theory with the B3LYP/6-31G* basis set, the relative energies of s- cis conformers of lutein were determined. All 512 s-cis conformers were calculated with PM3. A smaller, representative group was also studied using density functional theory. PM3 results were correlated systematically to B3LYP values and this enables the results to be calibrated. The relative energies of the conformers range from 1-30 kcal/mole, and many are dynamically accessible at normal temperatures. Four commercial formulations containing lutein were studied. The serum and macular pigment (MP) responses of human subjects to these lutein supplements with doses of 9 or 20 mg/day were measured, relative to a placebo, over a six month period. In each instance, lutein levels in serum increased and correlated with MP increases. The results demonstrate that responses are significantly dependent upon formulation and that components other than lutein have an important influence serum response.

Membrane associated transporter protein gene (SLC45A2) and the genetic basis of normal human pigmentation variation

This work is concerned with the genetic basis of normal human pigmentation variation. Specifically, the role of polymorphisms within the solute carrier family 45 member 2 (SLC45A2 or membrane associated transporter protein; MATP) gene were investigated with respect to variation in hair, skin and eye colour ― both between and within populations. SLC45A2 is an important regulator of melanin production and mutations in the gene underly the most recently identified form of oculocutaneous albinism. There is evidence to suggest that non-synonymous polymorphisms in SLC45A2 are associated with normal pigmentation variation between populations. Therefore, the underlying hypothesis of this thesis is that polymorphisms in SLC45A2 will alter the function or regulation of the protein, thereby altering the important role it plays in melanogenesis and providing a mechanism for normal pigmentation variation. In order to investigate the role that SLC45A2 polymorphisms play in human pigmentation variation, a DNA database was established which collected pigmentation phenotypic information and blood samples of more than 700 individuals. This database was used as the foundation for two association studies outlined in this thesis, the first of which involved genotyping two previously-described non-synonymous polymorphisms, p.Glu272Lys and p.Phe374Leu, in four different population groups. For both polymorphisms, allele frequencies were significantly different between population groups and the 272Lys and 374Leu alleles were strongly associated with black hair, brown eyes and olive skin colour in Caucasians. This was the first report to show that SLC45A2 polymorphisms were associated with normal human intra-population pigmentation variation. The second association study involved genotyping several SLC45A2 promoter polymorphisms to determine if they also played a role in pigmentation variation. Firstly, the transcription start site (TSS), and hence putative proximal promoter region, was identified using 5' RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE). Two alternate TSSs were identified and the putative promoter region was screened for novel polymorphisms using denaturing high performance liquid chromatography (dHPLC). A novel duplication (c.–1176_–1174dupAAT) was identified along with other previously described single nucleotide polymorphisms (c.–1721C>G and c.–1169G>A). Strong linkage disequilibrium ensured that all three polymorphisms were associated with skin colour such that the –1721G, +dup and –1169A alleles were associated with olive skin in Caucasians. No linkage disequilibrium was observed between the promoter and coding region polymorphisms, suggesting independent effects. The association analyses were complemented with functional data, showing that the –1721G, +dup and –1169A alleles significantly decreased SLC45A2 transcriptional activity. Based on in silico bioinformatic analysis that showed these alleles remove a microphthalmia-associated transcription factor (MITF) binding site, and that MITF is a known regulator of SLC45A2 (Baxter and Pavan, 2002; Du and Fisher, 2002), it was postulated that SLC45A2 promoter polymorphisms could contribute to the regulation of pigmentation by altering MITF binding affinity. Further characterisation of the SLC45A2 promoter was carried out using luciferase reporter assays to determine the transcriptional activity of different regions of the promoter. Five constructs were designed of increasing length and their promoter activity evaluated. Constitutive promoter activity was observed within the first ~200 bp and promoter activity increased as the construct size increased. The functional impact of the –1721G, +dup and –1169A alleles, which removed a MITF consensus binding site, were assessed using electrophoretic mobility shift assays (EMSA) and expression analysis of genotyped melanoblast and melanocyte cell lines. EMSA results confirmed that the promoter polymorphisms affected DNA-protein binding. Interestingly, however, the protein/s involved were not MITF, or at least MITF was not the protein directly binding to the DNA. In an effort to more thoroughly characterise the functional consequences of SLC45A2 promoter polymorphisms, the mRNA expression levels of SLC45A2 and MITF were determined in melanocyte/melanoblast cell lines. Based on SLC45A2’s role in processing and trafficking TYRP1 from the trans-Golgi network to stage 2 melanosmes, the mRNA expression of TYRP1 was also investigated. Expression results suggested a coordinated expression of pigmentation genes. This thesis has substantially contributed to the field of pigmentation by showing that SLC45A2 polymorphisms not only show allele frequency differences between population groups, but also contribute to normal pigmentation variation within a Caucasian population. In addition, promoter polymorphisms have been shown to have functional consequences for SLC45A2 transcription and the expression of other pigmentation genes. Combined, the data presented in this work supports the notion that SLC45A2 is an important contributor to normal pigmentation variation and should be the target of further research to elucidate its role in determining pigmentation phenotypes. Understanding SLC45A2’s function may lead to the development of therapeutic interventions for oculocutaneous albinism and other disorders of pigmentation. It may also help in our understanding of skin cancer susceptibility and evolutionary adaptation to different UV environments, and contribute to the forensic application of pigmentation phenotype prediction.

The concept of vulnerability and the protection of human subjects of research

This article provides an overview of the concept of vulnerability through the lens of the U.S. federal regulations for the protection of human subjects of research. General issues that emerge for nurse researchers working with regulated vulnerable populations are identified. Points of current controversy in the application of the regulations and current discourse about vulnerable groups are highlighted. Suggestions for negotiating the tension between federally regulated human subject requirements and the realities of research with vulnerable subjects are given. The limitations of the designation of vulnerable as a protection for human subjects will also be discussed.

Expected values for pedometer-determined physical activity in older populations

The purpose of this review is to update expected values for pedometer-determined physical activity in free-living healthy older populations. A search of the literature published since 2001 began with a keyword (pedometer, "step counter," "step activity monitor" or "accelerometer AND steps/day") search of PubMed, Cumulative Index to Nursing & Allied Health Literature (CINAHL), SportDiscus, and PsychInfo. An iterative process was then undertaken to abstract and verify studies of pedometer-determined physical activity (captured in terms of steps taken; distance only was not accepted) in free-living adult populations described as ≥ 50 years of age (studies that included samples which spanned this threshold were not included unless they provided at least some appropriately age-stratified data) and not specifically recruited based on any chronic disease or disability. We identified 28 studies representing at least 1,343 males and 3,098 females ranging in age from 50–94 years. Eighteen (or 64%) of the studies clearly identified using a Yamax pedometer model. Monitoring frames ranged from 3 days to 1 year; the modal length of time was 7 days (17 studies, or 61%). Mean pedometer-determined physical activity ranged from 2,015 steps/day to 8,938 steps/day. In those studies reporting such data, consistent patterns emerged: males generally took more steps/day than similarly aged females, steps/day decreased across study-specific age groupings, and BMI-defined normal weight individuals took more steps/day than overweight/obese older adults. The range of 2,000–9,000 steps/day likely reflects the true variability of physical activity behaviors in older populations. More explicit patterns, for example sex- and age-specific relationships, remain to be informed by future research endeavors.

Expected values for steps/day in special populations

Objective To assemble expected values for free-living steps/day in special populations living with chronic illnesses and disabilities. Method Studies identified since 2000 were categorized into similar illnesses and disabilities, capturing the original reference, sample descriptions, descriptions of instruments used (i.e., pedometers, piezoelectric pedometers, accelerometers), number of days worn, and mean and standard deviation of steps/day. Results Sixty unique studies represented: 1) heart and vascular diseases, 2) chronic obstructive lung disease, 3) diabetes and dialysis, 4) breast cancer, 5) neuromuscular diseases, 6) arthritis, joint replacement, and fibromyalgia, 7) disability (including mental retardation/intellectual difficulties), and 8) other special populations. A median steps/day was calculated for each category. Waist-mounted and ankle-mounted instruments were considered separately due to fundamental differences in assessment properties. For waist-mounted instruments, the lowest median values for steps/day are found in disabled older adults (1214 steps/day) followed by people living with COPD (2237 steps/day). The highest values were seen in individuals with Type 1 diabetes (8008 steps/day), mental retardation/intellectual disability (7787 steps/day), and HIV (7545 steps/day). Conclusion This review will be useful to researchers/practitioners who work with individuals living with chronic illness and disability and require such information for surveillance, screening, intervention, and program evaluation purposes. Keywords: Exercise; Walking; Ambulatory monitoring

Neural differentiation potential of human bone marrow-derived mesenchymal stromal cells: misleading marker gene expression

Background In contrast to pluripotent embryonic stem cells, adult stem cells have been considered to be multipotent, being somewhat more restricted in their differentiation capacity and only giving rise to cell types related to their tissue of origin. Several studies, however, have reported that bone marrow-derived mesenchymal stromal cells (MSCs) are capable of transdifferentiating to neural cell types, effectively crossing normal lineage restriction boundaries. Such reports have been based on the detection of neural-related proteins by the differentiated MSCs. In order to assess the potential of human adult MSCs to undergo true differentiation to a neural lineage and to determine the degree of homogeneity between donor samples, we have used RT-PCR and immunocytochemistry to investigate the basal expression of a range of neural related mRNAs and proteins in populations of non-differentiated MSCs obtained from 4 donors. Results The expression analysis revealed that several of the commonly used marker genes from other studies like nestin, Enolase2 and microtubule associated protein 1b (MAP1b) are already expressed by undifferentiated human MSCs. Furthermore, mRNA for some of the neural-related transcription factors, e.g. Engrailed-1 and Nurr1 were also strongly expressed. However, several other neural-related mRNAs (e.g. DRD2, enolase2, NFL and MBP) could be identified, but not in all donor samples. Similarly, synaptic vesicle-related mRNA, STX1A could only be detected in 2 of the 4 undifferentiated donor hMSC samples. More significantly, each donor sample revealed a unique expression pattern, demonstrating a significant variation of marker expression. Conclusion The present study highlights the existence of an inter-donor variability of expression of neural-related markers in human MSC samples that has not previously been described. This donor-related heterogeneity might influence the reproducibility of transdifferentiation protocols as well as contributing to the ongoing controversy about differentiation capacities of MSCs. Therefore, further studies need to consider the differences between donor samples prior to any treatment as well as the possibility of harvesting donor cells that may be inappropriate for transplantation strategies.

Interfacing Jack and anybody : towards anthropometric musculoskeletal digital human modeling

Virtual prototyping emerges as a new technology to replace existing physical prototypes for product evaluation, which are costly and time consuming to manufacture. Virtualization technology allows engineers and ergonomists to perform virtual builds and different ergonomic analyses on a product. Digital Human Modelling (DHM) software packages such as Siemens Jack, often integrate with CAD systems to provide a virtual environment which allows investigation of operator and product compatibility. Although the integration between DHM and CAD systems allows for the ergonomic analysis of anthropometric design, human musculoskeletal, multi-body modelling software packages such as the AnyBody Modelling System (AMS) are required to support physiologic design. They provide muscular force analysis, estimate human musculoskeletal strain and help address human comfort assessment. However, the independent characteristics of the modelling systems Jack and AMS constrain engineers and ergonomists in conducting a complete ergonomic analysis. AMS is a stand alone programming system without a capability to integrate into CAD environments. Jack is providing CAD integrated human-in-the-loop capability, but without considering musculoskeletal activity. Consequently, engineers and ergonomists need to perform many redundant tasks during product and process design. Besides, the existing biomechanical model in AMS uses a simplified estimation of body proportions, based on a segment mass ratio derived scaling approach. This is insufficient to represent user populations anthropometrically correct in AMS. In addition, sub-models are derived from different sources of morphologic data and are therefore anthropometrically inconsistent. Therefore, an interface between the biomechanical AMS and the virtual human model Jack was developed to integrate a musculoskeletal simulation with Jack posture modeling. This interface provides direct data exchange between the two man-models, based on a consistent data structure and common body model. The study assesses kinematic and biomechanical model characteristics of Jack and AMS, and defines an appropriate biomechanical model. The information content for interfacing the two systems is defined and a protocol is identified. The interface program is developed and implemented through Tcl and Jack-script(Python), and interacts with the AMS console application to operate AMS procedures.

A comprehensive literature review of the pelvis and the lower extremity digital human models under quasi-static conditions

Finite Element Modeling (FEM) has become a vital tool in the automotive design and development processes. FEM of the human body is a technique capable of estimating parameters that are difficult to measure in experimental studies with the human body segments being modeled as complex and dynamic entities. Several studies have been dedicated to attain close-to-real FEMs of the human body (Pankoke and Siefert 2007; Amann, Huschenbeth et al. 2009; ESI 2010). The aim of this paper is to identify and appraise the state of-the art models of the human body which incorporate detailed pelvis and/or lower extremity models. Six databases and search engines were used to obtain literature, and the search was limited to studies published in English since 2000. The initial search results identified 636 pelvis-related papers, 834 buttocks-related papers, 505 thigh-related papers, 927 femur-related papers, 2039 knee-related papers, 655 shank-related papers, 292 tibia-related papers, 110 fibula-related papers, 644 ankle related papers, and 5660 foot-related papers. A refined search returned 100 pelvis-related papers, 45 buttocks related papers, 65 thigh-related papers, 162 femur-related papers, 195 kneerelated papers, 37 shank-related papers, 80 tibia-related papers, 30 fibula-related papers and 102 ankle-related papers and 246 foot-related papers. The refined literature list was further restricted by appraisal against a modified LOW appraisal criteria. Studies with unclear methodologies, with a focus on populations with pathology or with sport related dynamic motion modeling were excluded. The final literature list included fifteen models and each was assessed against the percentile the model represents, the gender the model was based on, the human body segment/segments included in the model, the sample size used to develop the model, the source of geometric/anthropometric values used to develop the model, the posture the model represents and the finite element solver used for the model. The results of this literature review provide indication of bias in the available models towards 50th percentile male modeling with a notable concentration on the pelvis, femur and buttocks segments.

Floods and human health : a systematic review

Floods are the most common type of disaster globally, responsible for almost 53,000 deaths in the last decade alone (23:1 low- versus high-income countries). This review assessed recent epidemiological evidence on the impacts of floods on human health. Published articles (2004–2011) on the quantitative relationship between floods and health were systematically reviewed. 35 relevant epidemiological studies were identified. Health outcomes were categorized into short- and long-term and were found to depend on the flood characteristics and people's vulnerability. It was found that long-term health effects are currently not well understood. Mortality rates were found to increase by up to 50% in the first year post-flood. After floods, it was found there is an increased risk of disease outbreaks such as hepatitis E, gastrointestinal disease and leptospirosis, particularly in areas with poor hygiene and displaced populations. Psychological distress in survivors (prevalence 8.6% to 53% two years post-flood) can also exacerbate their physical illness. There is a need for effective policies to reduce and prevent flood-related morbidity and mortality. Such steps are contingent upon the improved understanding of potential health impacts of floods. Global trends in urbanization, burden of disease, malnutrition and maternal and child health must be better reflected in flood preparedness and mitigation programs.

ATF5, a possible regulator of osteogenic differentiation in human adipose-derived stem cells

The regulatory pathways involved in maintaining the pluripotency of embryonic stem cells are partially known, whereas the regulatory pathways governing adult stem cells and their "stem-ness" are characterized to an even lesser extent. We, therefore, screened the transcriptome profiles of 20 osteogenically induced adult human adipose-derived stem cell (ADSC) populations and investigated for putative transcription factors that could regulate the osteogenic differentiation of these ADSC. We studied a subgroup of donors' samples that had a disparate osteogenic response transcriptome from that of induced human fetal osteoblasts and the rest of the induced human ADSC samples. From our statistical analysis, we found activating transcription factor 5 (ATF5) to be significantly and consistently down-regulated in a randomized time-course study of osteogenically differentiated adipose-derived stem cells from human donor samples. Knockdown of ATF5 with siRNA showed an increased sensitivity to osteogenic induction. This evidence suggests a role for ATF5 in the regulation of osteogenic differentiation in adipose-derived stem cells. To our knowledge, this is the first report that indicates a novel role of transcription factors in regulating osteogenic differentiation in adult or tissue specific stem cells. © 2012 Wiley Periodicals, Inc.

Persistence of multiple genetic lineages within intra-host populations of Ross River virus

We examined the structure and extent of genetic diversity in intrahost populations of Ross River virus (RRV) in samples from six human patients, focusing on the nonstructural (nsP3) and structural (E2) protein genes. Strikingly, although the samples were collected from contrasting ecological settings 3,000 kilometers apart in Australia, we observed multiple viral lineages in four of the six individuals, which is indicative of widespread mixed infections. In addition, a comparison with previously published RRV sequences revealed that these distinct lineages have been in circulation for at least 5 years, and we were able to document their long-term persistence over extensive geographical distances

The wMel Wolbachia strain blocks dengue and invades caged Aedes aegypti populations

Dengue fever is the most important mosquito-borne viral disease of humans with more than 50 million cases estimated annually in more than 100 countries. Disturbingly, the geographic range of dengue is currently expanding and the severity of outbreaks is increasing. Control options for dengue are very limited and currently focus on reducing population abundance of the major mosquito vector, Aedes aegypti. These strategies are failing to reduce dengue incidence in tropical communities and there is an urgent need for effective alternatives. It has been proposed that endosymbiotic bacterial Wolbachia infections of insects might be used in novel strategies for dengue control. For example, the wMelPop-CLA Wolbachia strain reduces the lifespan of adult A. aegypti mosquitoes in stably transinfected lines. This life-shortening phenotype was predicted to reduce the potential for dengue transmission. The recent discovery that several Wolbachia infections, including wMelPop-CLA, can also directly influence the susceptibility of insects to infection with a range of insect and human pathogens has markedly changed the potential for Wolbachia infections to control human diseases. Here we describe the successful transinfection of A. aegypti with the avirulent wMel strain of Wolbachia, which induces the reproductive phenotype cytoplasmic incompatibility with minimal apparent fitness costs and high maternal transmission, providing optimal phenotypic effects for invasion. Under semi-field conditions, the wMel strain increased from an initial starting frequency of 0.65 to near fixation within a few generations, invading A. aegypti populations at an accelerated rate relative to trials with the wMelPop-CLA strain. We also show that wMel and wMelPop-CLA strains block transmission of dengue serotype 2 (DENV-2) in A. aegypti, forming the basis of a practical approach to dengue suppression.

Climate Change and Human Displacement

This paper will focus on the legal issues associated with people displaced as a result of water scarcity. Human displacement can lead to internal displacement (displacement of people within their country) and external displacement (displacement of people into another country). If the displacement takes place as a result of climate change these people may be referred to as climate refugees. The majority of work on climate refugees has focused on those people that will lose their homes as a result of sea –level rise. The number of people that could be displaced as a result of prolonged drought and lack of adequate water supplies is likely to be far more significant in number. There are estimates that around 2.8 billion people will suffer water shortages by 2025 and many of these people are at increased risk of internal or external displacement. Certain groups are more likely to be displaced as a result of prolonged drought or water scarcity. These groups include indigenous and minorities groups living in areas that are more susceptible to climate change and groups living in areas with a history of water shortage and supply issues. People displaced as a result of water scarcity are at increased risks of malnutrition and of dehydration. Furthermore the lack of adequate water supplies in such areas increases the risk and spread of disease among the population. In certain instances internal and external displacement may lead to escalation of conflict and competition for water resources in newly settled territories. This paper will use case studies from Australia (indigenous groups and rural landholders) and East Africa (Ethiopia, Sudan and Kenya) to demonstrate the significance of human displacement arising as a result of water scarcity. Climate adaptation policy frameworks will need to address a number of legal issues, arising as a result of climate displacement from water scarcity. There are a number of unresolved legal issues for both categories of environmental displaced people. The major legal issue for externally environmentally displaced people is lack of international recognition and support for these people. The Climate Change Convention, the Refugee Convention, the Desertification Convention and Human Rights instruments all fail to provide recognition for people externally displaced as a result of environmental conditions. Similarly there is a lack of legal recognition and legal support mechanisms to assist those people internally displaced by environmental conditions. The lack of developed environmental rights in most countries contributes to this problem. Polices and governance frameworks must be put in place which aims to prevent such displacement through programs identifying populations at risk and instigating damage mitigation and relocation programs. In addition there are a number of legal issues which may arise such as; rights of compensation, property and tenure disputes, increases on the water demand and environmental degradation in places of relocation and jurisdictional issues arising in federal countries. This paper will provide an overview of the legal issues at the international and national levels arising as a result of climate displacement from water scarcity.  

Evolutionary genetics and human assisted movement of a globally invasive pest (Russian wheat aphid : Diuraphis noxia)

This PhD study has examined the population genetics of the Russian wheat aphid (RWA, Diuraphis noxia), one of the world’s most invasive agricultural pests, throughout its native and introduced global range. Firstly, this study investigated the geographic distribution of genetic diversity within and among RWA populations in western China. Analysis of mitochondrial data from 18 sites provided evidence for the long-term existence and expansion of RWAs in western China. The results refute the hypothesis that RWA is an exotic species only present in China since 1975. The estimated date of RWA expansion throughout western China coincides with the debut of wheat domestication and cultivation practices in western Asia in the Holocene. It is concluded that western China represents the limit of the far eastern native range of this species. Analysis of microsatellite data indicated high contemporary gene flow among northern populations in western China, while clear geographic isolation between northern and southern populations was identified across the Tianshan mountain range and extensive desert regions. Secondly, this study analyzed the worldwide pathway of invasion using both microsatellite and endosymbiont genetic data. Individual RWAs were obtained from native populations in Central Asia and the Middle East and invasive populations in Africa and the Americas. Results indicated two pathways of RWA invasion from 1) Syria in the Middle East to North Africa and 2) Turkey to South Africa, Mexico and then North and South America. Very little clone diversity was identified among invasive populations suggesting that a limited founder event occurred together with predominantly asexual reproduction and rapid population expansion. The most likely explanation for the rapid spread (within two years) from South Africa to the New World is by human movement, probably as a result of the transfer of wheat breeding material. Furthermore, the mitochondrial data revealed the presence of a universal haplotype and it is proposed that this haplotype is representative of a wheat associated super-clone that has gained dominance worldwide as a result of the widespread planting of domesticated wheat. Finally, this study examined salivary gland gene diversity to determine whether a functional basis for RWA invasiveness could be identified. Peroxidase DNA sequence data were obtained for a selection of worldwide RWA samples. Results demonstrated that most native populations were polymorphic while invasive populations were monomorphic, supporting previous conclusions relating to demographic founder effects in invasive populations. Purifying selection most likely explains the existence of a universal allele present in Middle Eastern populations, while balancing selection was evident in East Asian populations. Selection acting on the peroxidase gene may provide an allele-dependent advantage linked to the successful establishment of RWAs on wheat, and ultimately their invasion potential. In conclusion, this study is the most comprehensive molecular genetic investigation of RWA population genetics undertaken to date and provides significant insights into the source and pathway of global invasion and the potential existence of a wheat-adapted genotype that has colonised major wheat growing countries worldwide except for Australia. This research has major biosecurity implications for Australia’s grain industry.

A city for all citizens : integrating children and youth from marginalized populations into city planning

Socially just, intergenerational urban spaces should not only accommodate children and adolescents, but engage them as participants in the planning and design of welcoming spaces. With this goal, city agencies in Boulder, Colorado, the Boulder Valley School District, the Children, Youth and Environments Center at the University of Colorado, and a number of community organizations have been working in partnership to integrate young people’s ideas and concerns into the redesign of parks and civic areas and the identification of issues for city planning. Underlying their work is a commitment to the Convention on the Rights of the Child and children’s rights to active citizenship from a young age. This paper describes approaches used to engage with young people and methods of participation, and reflects on lessons learned about how to most effectively involve youth from underrepresented populations and embed diverse youth voices into the culture of city planning.