Bioinformatics mining for disease causing mutations

Humans and dogs share many common diseases, and it has been shown that the identification of mutations that cause disease in dogs can help unravel the genetic basis for a similar disease in humans. Mapping of traits and disease in dogs is not a new idea, but the sequencing of the whole dog genome, the creation of a dense SNP maps followed by the development of SNP arrays for high throughput genotyping has led to new facilitated mapping procedures. Each dog breed can be seen as a genetic isolate and certain breeds are often predisposed to specific diseases. Because of the genomic structure of the dog genome and the availability of new resources for disease mapping, the dog has been proposed to be especially advantageous for the mapping of complex disease that is difficult to map in human outbred populations. In this thesis, the aim has been to identify disease-causing mutations for three complex diseases in dogs with the presence of similar conditions in humans. Emphasis has been on bioinformatics analyses of genome-wide SNP and large re-sequencing data. In the dog breed Nova Scotia duck tolling retriever it is common with an immune-mediated disease complex that resembles human systemic lupus erythematosus (SLE). In paper I we used a two-stage genome-wide association mapping method and successfully located several susceptibility loci in dogs for this disease complex. In paper II we identified a mutation that had been under selection in the Shar-Pei breed, causing both a breed-defining wrinkled skin phenotype and an autoinflammatory fever disease. Because the locus had been under selection we used an alternative mapping approach, called homozygosity mapping to identify the locus, followed by re-sequencing using next generation sequencing technologies. In paper III we report the development of a web-based tool that facilitates analyses and extraction of essential information from the large amount of data produced by next generation sequencing projects. In paper IV we used across-breed genome-wide association mapping to identify risk factors for glioma, a type of malignant brain tumor fatal to both human and dogs. For the three diseases excellent candidate genes have been identified, and continued research might has the potential to lead to better treatment options and thus benefit both dogs and humans

Localization of Canine Brachycephaly Using an Across Breed Mapping Approach

The domestic dog, Canis familiaris, exhibits profound phenotypic diversity and is an ideal model organism for the genetic dissection of simple and complex traits. However, some of the most interesting phenotypes are fixed in particular breeds and are therefore less tractable to genetic analysis using classical segregation-based mapping approaches. We implemented an across breed mapping approach using a moderately dense SNP array, a low number of animals and breeds carefully selected for the phenotypes of interest to identify genetic variants responsible for breed-defining characteristics. Using a modest number of affected (10–30) and control (20–60) samples from multiple breeds, the correct chromosomal assignment was identified in a proof of concept experiment using three previously defined loci; hyperuricosuria, white spotting and chondrodysplasia. Genome-wide association was performed in a similar manner for one of the most striking morphological traits in dogs: brachycephalic head type. Although candidate gene approaches based on comparable phenotypes in mice and humans have been utilized for this trait, the causative gene has remained elusive using this method. Samples from nine affected breeds and thirteen control breeds identified strong genome-wide associations for brachycephalic head type on Cfa 1. Two independent datasets identified the same genomic region. Levels of relative heterozygosity in the associated region indicate that it has been subjected to a selective sweep, consistent with it being a breed defining morphological characteristic. Genotyping additional dogs in the region confirmed the association. To date, the genetic structure of dog breeds has primarily been exploited for genome wide association for segregating traits. These results demonstrate that non-segregating traits under strong selection are equally tractable to genetic analysis using small sample numbers

A Frameshift Mutation in Golden Retriever Dogs with Progressive Retinal Atrophy Endorses SLC4A3 as a Candidate Gene for Human Retinal Degenerations

Progressive retinal atrophy (PRA) in dogs, the canine equivalent of retinitis pigmentosa (RP) in humans, is characterised by vision loss due to degeneration of the photoreceptor cells in the retina, eventually leading to complete blindness. It affects more than 100 dog breeds, and is caused by numerous mutations. RP affects 1 in 4000 people in the Western world and 70% of causal mutations remain unknown. Canine diseases are natural models for the study of human diseases and are becoming increasingly useful for the development of therapies in humans. One variant, prcd-PRA, only accounts for a small proportion of PRA cases in the Golden Retriever (GR) breed. Using genome-wide association with 27 cases and 19 controls we identified a novel PRA locus on CFA37 (praw = 1.94×10−10, pgenome = 1.0×10−5), where a 644 kb region was homozygous within cases. A frameshift mutation was identified in a solute carrier anion exchanger gene (SLC4A3) located within this region. This variant was present in 56% of PRA cases and 87% of obligate carriers, and displayed a recessive mode of inheritance with full penetrance within those lineages in which it segregated. Allele frequencies are approximately 4% in the UK, 6% in Sweden and 2% in France, but the variant has not been found in GRs from the US. A large proportion of cases (approximately 44%) remain unexplained, indicating that PRA in this breed is genetically heterogeneous and caused by at least three mutations. SLC4A3 is important for retinal function and has not previously been associated with spontaneously occurring retinal degenerations in any other species, including humans

A Novel Unstable Duplication Upstream of HAS2 Predisposes to a Breed-Defining Skin Phenotype and a Periodic Fever Syndrome in Chinese Shar-Pei Dogs

Hereditary periodic fever syndromes are characterized by recurrent episodes of fever and inflammation with no known pathogenic or autoimmune cause. In humans, several genes have been implicated in this group of diseases, but the majority of cases remain unexplained. A similar periodic fever syndrome is relatively frequent in the Chinese Shar-Pei breed of dogs. In the western world, Shar-Pei have been strongly selected for a distinctive thick and heavily folded skin. In this study, a mutation affecting both these traits was identified. Using genome-wide SNP analysis of Shar-Pei and other breeds, the strongest signal of a breed-specific selective sweep was located on chromosome 13. The same region also harbored the strongest genome-wide association (GWA) signal for susceptibility to the periodic fever syndrome (praw = 2.3×10−6, pgenome = 0.01). Dense targeted resequencing revealed two partially overlapping duplications, 14.3 Kb and 16.1 Kb in size, unique to Shar-Pei and upstream of the Hyaluronic Acid Synthase 2 (HAS2) gene. HAS2 encodes the rate-limiting enzyme synthesizing hyaluronan (HA), a major component of the skin. HA is up-regulated and accumulates in the thickened skin of Shar-Pei. A high copy number of the 16.1 Kb duplication was associated with an increased expression of HAS2 as well as the periodic fever syndrome (p<0.0001). When fragmented, HA can act as a trigger of the innate immune system and stimulate sterile fever and inflammation. The strong selection for the skin phenotype therefore appears to enrich for a pleiotropic mutation predisposing these dogs to a periodic fever syndrome. The identification of HA as a major risk factor for this canine disease raises the potential of this glycosaminoglycan as a risk factor for human periodic fevers and as an important driver of chronic inflammation

Ökning av svarsfrekvens och påskyndning av återlämning för hälsoenkäter : - En studie vid Danderyds sjukhus högspecialiserade smärtmottagning

Health questionnaires are widely used. A low response rate can lead to biased results and affect the validity of the study. This thesis investigates what can be done to increase the response rate and expedite the return of health questionnaires. The main aim of the study is to provide suggestions on how to improve the response rate to the National Pain Registry’s questionnaire at Danderyd Specialized Pain Rehab Clinic, although the result is significant in other contexts where questionnaires are used and a high response rate desired. Material has been collected through literature search, observations, interviews with patients at Danderyd Specialized Pain Rehab Clinic and interviews with clinics that have a high response rate according to the National Pain Registry’s reports. Several factors affecting response rate was found in the literature studies and interviews. Providing a digitalized alternative, clearly explaining the aim of the study and why completing the questionnaire is important and making the participants feel like stakeholders in the study are examples of areas where improvements can be made. According to the patient interviews about the National Pain Registries first and second questionnaire some problems to fill in the questionnaires were due to difficulties in estimating the pain because of shifting pain intensity, unclear phrasing of the questions or answer and a feeling of lack of someone to ask. Reasons for not returning the one-year follow-up questionnaire were that the questionnaire never was received, it got cluttered and therefore never mailed back or that it was not seen as important nor beneficial to the patient. Suggestions made to increase response rate and expedite the return of the questionnaires based on the findings include increased information about the questionnaires and the National Pain Registry, notifications and reminders to help patients remember and tracking the mailed questionnaire to make sure it reaches the patient. Changing the front-page layout could make it more interesting to look at. To clarify questions, the option to complete all questionnaires at the clinic could be given and a computer alternative. The option to hand back the questionnaire either by mail, digitally or at the visit along with a scheduled one-year follow-up meeting could both increase response rate and expedite the return.  Hälsoenkäter används omfattande. En låg svarsfrekvens kan leda till missvisande resultat och påverka giltigheten av studien. Det här examensarbetet undersöker vad som kan göras för att öka svarsfrekvensen och påskynda återlämningen av hälsoenkäter. Det huvudsakliga målet med studien är att ge förslag som kan öka svarsfrekvensen till det Nationella Registret över Smärtrehabiliterings frågeformulär vid Danderyds Högspecialiserade Smärtmottagning, även om resultatet är betydelsefullt även i andra sammanhang där frågeformulär används och en hög svarsfrekvens önskas. Material har samlats in genom litteraturstudier, observationer, intervjuer med patienter vid Danderyds Högspecialiserade Smärtmottagning och intervjuer med kliniker som har en hög svarsfrekvens enligt det Nationella Registret över Smärtrehabiliterings rapporter. Flertalet faktorer som påverkar svarsfrekvens enligt litteraturstudier och intervjuerna hittades. Tillhandahålla ett digitalt alternativ, klargöra och förklara målet med studien och varför formuläret är viktigt att fylla i och att få deltagarna att känna sig som intressenter i studien är exempel på områden där förbättringar kan ske. Enligt patientintervjuerna angående nationella smärtrehabiliteringens första och andra frågeformuläret beror en del av problemen att fylla i formuläret på att smärtan skiftar i intensitet, oklara formuleringar av frågor och svar och en känsla av brist på någon att fråga. Skäl till att ett-årsuppföljningsformuläret inte återlämnades var att det aldrig mottogs, att det blev plottrigt och därför aldrig skickades tillbaka eller att det inte sågs som varken viktigt eller fördelaktigt att fylla i av patienten. Förslag på hur svarsfrekvensen kan ökas och återlämning påskyndas av frågeformulären baserat på upptäckterna inkluderar ökad information om frågeformulären och det Nationella Registret över Smärtrehabilitering, notifieringar och påminnelser för att hjälpa patienterna att komma ihåg och spårning av brevet med formuläret för att se till att det når patienten. Ändring av framsidans utseende skulle kunna göra det intressantare att titta på. För att klargöra frågor skulle en valmöjlighet att fylla i alla formulären på kliniken kunna ges samt ett datoralternativ. En valmöjlighet att ge tillbaka formuläret antingen via brev, digitalt eller för hand tillsammans med ett schemalagt ett-årsuppföljningsmöte kan både öka svarsfrekvensen och påskynda återlämningen av formuläret

Using combined methods to reveal the dynamic organization of protein networks

Proteins combine in various ways to execute different essential functions. Cellular processes are enormously complex and it is a great challenge to explain the underlying organization. Various methods have been applied in attempt to reveal the organization of the cell. Gene expression analysis uses the mRNA levels in the cell to predict which proteins are present in the cell simultaneously. This method is useful but also known to sometimes fail. Proteins that are known to be functionally related do not always show a significant correlation in gene expression. This fact might be explained by the dynamic organization of the proteome. Proteins can have diverse functions and might interact with some proteins only during a few time points, which would probably not result in significant correlation in their gene expression. In this work we tried to address this problem by combining gene expression data with data for physical interactions between proteins. We used a method for modular decomposition introduced by Gagneur et al. (2004) that aims to reveal the logical organization in protein-protein networks. We extended the interpretation of the modular decomposition to localize the dynamics in the protein organization. We found evidence that protein-interactions supported by gene expression data are very likely to be related in function and thus can be used to predict function for unknown proteins. We also identified negative correlation in gene expression as an overlooked area. Several hypotheses were generated using combination of these methods. Some could be verified by the literature and others might shed light on new pathways after additional experimental testing

Ökning av svarsfrekvens och påskyndning av återlämning för hälsoenkäter : - En studie vid Danderyds sjukhus högspecialiserade smärtmottagning

Health questionnaires are widely used. A low response rate can lead to biased results and affect the validity of the study. This thesis investigates what can be done to increase the response rate and expedite the return of health questionnaires. The main aim of the study is to provide suggestions on how to improve the response rate to the National Pain Registry’s questionnaire at Danderyd Specialized Pain Rehab Clinic, although the result is significant in other contexts where questionnaires are used and a high response rate desired. Material has been collected through literature search, observations, interviews with patients at Danderyd Specialized Pain Rehab Clinic and interviews with clinics that have a high response rate according to the National Pain Registry’s reports. Several factors affecting response rate was found in the literature studies and interviews. Providing a digitalized alternative, clearly explaining the aim of the study and why completing the questionnaire is important and making the participants feel like stakeholders in the study are examples of areas where improvements can be made. According to the patient interviews about the National Pain Registries first and second questionnaire some problems to fill in the questionnaires were due to difficulties in estimating the pain because of shifting pain intensity, unclear phrasing of the questions or answer and a feeling of lack of someone to ask. Reasons for not returning the one-year follow-up questionnaire were that the questionnaire never was received, it got cluttered and therefore never mailed back or that it was not seen as important nor beneficial to the patient. Suggestions made to increase response rate and expedite the return of the questionnaires based on the findings include increased information about the questionnaires and the National Pain Registry, notifications and reminders to help patients remember and tracking the mailed questionnaire to make sure it reaches the patient. Changing the front-page layout could make it more interesting to look at. To clarify questions, the option to complete all questionnaires at the clinic could be given and a computer alternative. The option to hand back the questionnaire either by mail, digitally or at the visit along with a scheduled one-year follow-up meeting could both increase response rate and expedite the return.  Hälsoenkäter används omfattande. En låg svarsfrekvens kan leda till missvisande resultat och påverka giltigheten av studien. Det här examensarbetet undersöker vad som kan göras för att öka svarsfrekvensen och påskynda återlämningen av hälsoenkäter. Det huvudsakliga målet med studien är att ge förslag som kan öka svarsfrekvensen till det Nationella Registret över Smärtrehabiliterings frågeformulär vid Danderyds Högspecialiserade Smärtmottagning, även om resultatet är betydelsefullt även i andra sammanhang där frågeformulär används och en hög svarsfrekvens önskas. Material har samlats in genom litteraturstudier, observationer, intervjuer med patienter vid Danderyds Högspecialiserade Smärtmottagning och intervjuer med kliniker som har en hög svarsfrekvens enligt det Nationella Registret över Smärtrehabiliterings rapporter. Flertalet faktorer som påverkar svarsfrekvens enligt litteraturstudier och intervjuerna hittades. Tillhandahålla ett digitalt alternativ, klargöra och förklara målet med studien och varför formuläret är viktigt att fylla i och att få deltagarna att känna sig som intressenter i studien är exempel på områden där förbättringar kan ske. Enligt patientintervjuerna angående nationella smärtrehabiliteringens första och andra frågeformuläret beror en del av problemen att fylla i formuläret på att smärtan skiftar i intensitet, oklara formuleringar av frågor och svar och en känsla av brist på någon att fråga. Skäl till att ett-årsuppföljningsformuläret inte återlämnades var att det aldrig mottogs, att det blev plottrigt och därför aldrig skickades tillbaka eller att det inte sågs som varken viktigt eller fördelaktigt att fylla i av patienten. Förslag på hur svarsfrekvensen kan ökas och återlämning påskyndas av frågeformulären baserat på upptäckterna inkluderar ökad information om frågeformulären och det Nationella Registret över Smärtrehabilitering, notifieringar och påminnelser för att hjälpa patienterna att komma ihåg och spårning av brevet med formuläret för att se till att det når patienten. Ändring av framsidans utseende skulle kunna göra det intressantare att titta på. För att klargöra frågor skulle en valmöjlighet att fylla i alla formulären på kliniken kunna ges samt ett datoralternativ. En valmöjlighet att ge tillbaka formuläret antingen via brev, digitalt eller för hand tillsammans med ett schemalagt ett-årsuppföljningsmöte kan både öka svarsfrekvensen och påskynda återlämningen av formuläret

Nya tillämpningar av Antrad Medicals upptiningsteknik : Klinik- och laboratorietillämpningar

Antrad Medical has developed an ultra-fast blood plasma thawing device named UFT100. The method is based on thawing with an oscillating electrical field and unlike water baths it is a dry method. Fast and homogeneous thawing is achieved. This project investigates new possible applications where this thawing technology could be used within the clinical and laboratory segment. The aim was to identify existing thawing and heating methods for a substance that can be improved and potentially replaced with the UFT100.   Data has been collected through literature research and interviews with Antrad Medical and specialists working in Sweden. The specialists are working within the clinical and laboratory field. A number of criteria for establishment of the UFT100 were set up and used as a tool for evaluation.   The substances investigated during this project were infusion medicine, embryos, substance in a sampling tube, protein based pharmaceuticals, stem cells for transplantation and research, biobank sampling tubes, cryoprecipitate, human hepatocytes, live vaccines, API, BDS, intermediate and antibodies. Two applications within the clinical area are found probable, stem cells and cryoprecipitate. Further investigation for human hepatocytes, API, BDS, intermediate and pharmaceuticals is needed.Antrad Medical har utvecklat en ultra-snabb blodplasmaupptinare kallad UFT100. Det är till skillnad från vattenbad en torr metod baserad på upptining med hjälp av oscillerande elektriska fält. Genom detta uppnås snabb och homogen upptining. Detta projekt undersöker nya möjliga kliniska- och laboratorietillämpningar för upptiningstekniken. Målet var att identifiera substanser vars nuvarande upptining- och uppvärmningsteknik kan förbättras och kanske ersättas med UFT100. Data har samlats in genom litteratursökning och genom intervjuer med Antrad Medical och specialister som arbetar i Sverige. Specialisterna arbetar inom kliniska områden och laboratorieverksamheter. Ett antal kriterier för etablering av UFT100 sattes upp och användes för utvärdering. Substanserna som undersöktes under projektets gång var infusionsmedicin, embryon, innehåll i ett provrör, proteinbaserade läkemedel, stamceller för transplantation och forskning, biobank-provrör, kryoprecipitat, humana hepatocyter, levande vaccin, aktiva substanser, läkemedelssubstanser, intermediat och antikroppar. Två tillämpningar inom det kliniska området ses som möjliga, stamceller och kryoprecipitat. Humana hepatocyter, aktiva substanser, läkemedelssubstanser och intermediat behöver undersökas vidare

Exploring Terra Incognita in the Design Space of Query Devices

This paper introduces query devices, an extension to the widget concept. Query devices are graphical database interaction objects such as rangesliders and toggles. The design space of query devices is structured and it is shown how the design space can be explored so that consistent and distinguishable query devices are created. Existing query devices are placed in the design space and new ones suggested. A set of query devices aimed for dynamic queries systems is presented