<p>Vitamin E is a natural lipophilic antioxidant contains eight structurally related forms, i.e., α-, β-, γ-, δ-tocopherols (αT, βT, γT, and δT) and corresponding tocotrienols. Recent research indicates that vitamin E forms are differentially metabolized to various carboxychromanols. Some these vitamin E metabolites have been shown to exhibit strong anti-inflammatory and anticancer effects, yet little is known about their bioavailability. Without this knowledge, it is impossible to assess the role of vitamin E metabolism in biological functions of vitamin E forms and their protective effects on chronic diseases. While αT and γT appear to improved gut health, the underlying mechanisms are not well understood. Furthermore, specific forms of vitamin E such as γT have been reported to have cancer-preventing effects, but their anticancer efficacy is relatively modest. For these reasons, this dissertation focused on the characterization of the pharmacokinetic formation of vitamin E metabolites after supplementation, and the investigation of the underlying mechanisms of the protective effect of vitamin E forms, αT and γT, on gut health, as well as anticancer efficacy of the combination of aspirin and γT on carcinogen-induced colon tumorigenesis.</p><p><br></p><p>The first project focuses on characterizing the pharmacokinetic formation of vitamin E metabolites after single dose supplementation of γ-tocopherol-rich mixed tocopherol (γTmT) and δ-tocotrienol (δTE). With our recently developed LC/MS/MS assay for quantifying vitamin E metabolites, we can simultaneously quantify the level of short-chain, long-chain, and sulfated carboxychromanols in plasma, urine, and fecal samples of supplemented animals. In this study, we investigated the pharmacokinetics including excretion of vitamin E forms and the formation of their metabolites after a single dose intragastric administration of tocopherols and tocotrienols in rats. We also measured vitamin E metabolites in the serum obtained from healthy humans after gT supplementation. In the plasma of rat, the pharmacokinetic profiles of γT and δTE are described as the following: γT, Cmax = 25.6 ± 9.1 μM, Tmax = 4 h; δTE, Cmax = 16.0 ± 2.3 μM, Tmax = 2 h. Sulfated CEHCs and sulfated 11’-COOHs were the predominant metabolites in the plasma of rat with Cmax of 0.4-0.5 μM (Tmax ~ 5-7 h) or ~0.3 μM (Tmax at 4.7 h), respectively. In 24-h urine, 2.7% of γT and 0.7% of dTE were excreted as conjugated CEHCs, the major identified urinary metabolites. In the feces, 17-45% of supplemented vitamers were excreted as un-metabolized forms and 4.9-9.2% as metabolites. The majority of metabolites excreted in feces were unconjugated carboxychromanols, among which 13’-COOHs constituted ~50% of total metabolites. Interestingly, 13’-COOHs derived from δTE were 2-fold higher than 13’-COOH from γT. Unlike rats, γ-CEHC is the predominant metabolites found in human plasma, although 11’-COOHs and 13’-COOHs (sulfated and unconjugated) were elevated by >20 folds responding to γT supplement. In this study, we found that tocopherols and tocotrienols, when taken as supplements, are mainly excreted as un-metabolized forms and long-chain carboxychromanols in feces. High fecal availability of 13’-COOHs may contribute to modulating effects on gut health.</p><p><br></p><p>The second project of my dissertation investigated the effect of vitamin E forms, αT and γT, on intestinal barrier function in a cellular model and a mouse colitis model. Inflammatory bowel diseases (IBD) are chronic idiopathic inflammatory conditions characterized by disruption of intestinal barrier integrity. Previous studies by others and us had demonstrated that vitamin E forms, αT and γT, can protect against chemical-induced colitis in animal models. However, the role of these vitamin E forms on intestinal barrier function has not been studied. Herein, we investigated the potential protective effects of vitamin E forms, αT and γT, on intestinal barrier function in a Caco-2 colon epithelial cell model and a dextran sodium sulfate (DSS)-induced colitis mouse model. In Caco-2 cells, pretreatment with 25mM αT and γT attenuated Caco-2 monolayer barrier dysfunction induced by 10 ng/mL TNF-α/IFN-γ, suggesting that these vitamin E forms protect intestinal barrier integrity in this cellular model. In male BALB/c mice, the supplementation of αT (0.05%) or γTmT (0.05%) when given 3 weeks before DSS treatment or at the same time as DSS treatment alleviated DSS-induced fecal bleeding and diarrhea symptoms in mice, and attenuated colon inflammation and colitis-associated damages. Additionally, αT and γTmT supplementation attenuated DSS-induced intestinal barrier dysfunction, as indicated by improving the level of occludin, a tight junction protein, in the colon and reducing lipopolysaccharide-binding protein (LBP) in the plasma. Furthermore, gut microbiota analysis demonstrated that αT and γTmT supplementation could modulate intestinal microbiome composition in mice with DSS treatment. DSS treatment reduced the relative abundance of Lachnospiraceae compared to healthy mice, and supplementation of αT and γT partially reversed this effect. Interestingly, the family Lachnospiraceae has been reported to decrease in IBD patients. Our study demonstrated the protective effects of vitamin E forms on intestinal barrier integrity in a cell-based model and a colitis model in mice. Furthermore, we demonstrated that these vitamin E forms caused favorable changes in the intestinal microbial population under colitis condition.</p><p><br></p><p>The third project of my dissertation evaluated the anticancer efficacy of the combination of aspirin and γT using an azoxymethane (AOM)-induced and colitis-promoted colon tumorigenesis mouse model. Extensive inflammation in the colon promotes the development of colorectal cancer (CRC). Eicosanoid production by pro-inflammatory enzymes, cyclooxygenases (COX-1 and COX-2) and 5-lipoxygenase (5-LOX) play a critical role in the initiation, progression, and invasion of CRC. Thus, nonsteroidal anti-inflammatory drugs (NSAIDs), such as aspirin, have been recommended for chemoprevention of CRC. However, long-term use of aspirin can cause many side effects, and the anticancer activity of aspirin is very modest. Previously, we have demonstrated that the combination of γT with aspirin prolonged the anti-inflammatory activity of aspirin and alleviated aspirin-associated adverse effects in a carrageenan-induced inflammation model in rats. Additionally, we found that the combination of γT and aspirin has stronger anticancer activity than aspirin or γT alone against HCT-116 human colorectal carcinoma cells. Therefore, we examined the anticancer effect of the combination of 0.025% aspirin and 0.05% γT against AOM-induced and DSS-promoted tumorigenesis in mice. In this study, we have found that the combination of aspirin and γT, but not aspirin or γT alone, suppressed colon tumorigenesis in mice, as indicated by 40% and 50% reduction in the multiplicity of total polyps (P < 0.05) and large adenomatous polyps (>2mm2, P < 0.05), respectively. More strikingly, the combination of aspirin and γT reduced the overall tumor area by 60% (P < 0.05). Noteworthy, the supplementation of γT also alleviated aspirin-induced stomach lesion and appeared to modulate intestinal microbial composition. Our study demonstrated that the combination of aspirin and γT has stronger anticancer activity than aspirin or γT alone while alleviates aspirin-associated adverse effect, suggesting that the combination of γT and aspirin is a more effective and safer chemopreventive agent for CRC than aspirin alone.</p>

Indiana University-Purdue University Indianapolis (IUPUI) / Non-alcoholic fatty liver disease and metabolic syndrome induced by high nutrient status have increasingly become a global health concern as it cause multiple complications. The mTOR complex is central in regulating anabolic reactions within cells under growth factors or under high nutrients stimulation. Constitutive and persistent activation of mTOR can impair cellular functions. In the first part of this study, we demonstrate a damping oscillation of mTOR activity during a long-term treatment of high fat diet. TFEB translocation and lysosomal enzyme activity also oscillate, but in an opposite direction. TFEB controls the lysosomal activity, autophagic degradation and lipid metabolism. Overexpression of wild type and mutant TFEB could inhibit NAFLD development in mice. In addition, TFEB location in nucleus inversely correlates with NAFLD severity in patients. mTOR activation under hypernutrition status suppresses TFEB translocation, inhibits lysosomal functions and autophagic degradation of lipid droplets. Inhibition of mTOR activity by rapamycin reverse the above phenotypes. Because mTOR activation also requires normal lysosomal function, the inhibition of TFEB by mTOR leads to decreased lysosomal function and mTOR downregulation. This negative feedback may explain the oscillation pattern of mTOR activation in long term high fat diet regimen and is a novel mechanism for inhibition of mTOR. In the second part of study, we report that Bid protein, previously known for its pro-apoptosis function in promoting mitochondrial permeability, plays an unexpected role in regulating fatty acid beta oxidation. Deletion of Bid in mice reprograms the body's response to hyper-nutrition caused by high fat diet, leading to the resistance to the development of obesity, liver steatosis and metabolic syndrome. These mice present a higher oxygen consumption, a lower respiratory quotient, and an increased beta-oxidation rate. Mechanistically, the high fat diet regimen triggers translocation of the full length Bid molecule to mitochondrial membrane. Genetic deletion of Bid also affects the stability of its binding protein, MTCH2 in the mitochondrial membrane. In summary, we describe in this study a mTOR-TFEB-lysosome feedback loop, which can regulate NAFLD development, and a novel Bid-mediated regulatory mechanism in beta-oxidation, which limits energy expenditure and promotes obesity development.

Primary sclerosing cholangitis (PSC) is a rare inflammatory chronic liver disease that causes damage to the intra- and or extrahepatic bile ducts leading to cholestasis. As the disease proceeds the development of cirrhosis and eventually liver failure occurs. This study aims to determine the role of IgG subclasses in the prognosis of PSC and its outcome. A retrospective analysis was performed of 183 patients followed at the Department of Upper Abdominal Diseases at the Karolinska University Hospital. Factors that were analysed were sex, age at PSC diagnosis, total IgG values, IgG subclasses values and events of autoimmune hepatitis (AIH), inflammatory bowel disease (IBD), colectomy, cirrhosis, cholangitis, endoscopic retrograde cholangiopancreatography (ERCP), liver transplantation and cholangiocarcinoma. This study showed that high IgG4 levels were associated with a higher incidence of cirrhosis, liver transplantation, cholangitis and ERCP, while low IgG4 levels were associated with a prior IBD diagnosis. In conclusion, elevated IgG4 levels were associated with a higher occurrence of cirrhosis, cholangitis, ERCP and liver transplantation. It seems that IgG4 could be of importance for outcome prediction in PSC.

First identified over a decade ago, Astrocyte Elevated Gene-1 (AEG-1) has been studied extensively due to early reports of its overexpression in various cancer cell lines. Research groups all over the globe including our own have since identified AEG-1 overexpression in cancers of diverse lineages including cancers of the liver, colon, skin, prostate, breast, lung, esophagus, neurons and neuronal glia as compared to matched normal tissue. A comprehensive and convincing body of data currently points to AEG-1 as an essential component, critical to the progression and perhaps onset of cancer. AEG-1 is a potent activator of multiple pro-tumorigenic signal transduction pathways such as mitogen-activated protein extracellular kinase (MEK)/ extracellular signal-regulated kinase (ERK), phosphotidyl-inositol-3-kinase (PI3K)/Akt/mTOR, NF-κB and Wnt/β-catenin pathway. In addition, studies show that AEG-1 not only alters global gene and protein expression profiles, it also modulates fundamental intracellular processes, such as transcription, translation and RNA interference in cancer cells most likely by functioning as a scaffold protein. The mechanisms by which AEG-1 is overexpressed in cancer have been studied extensively and it is clear that multiple layers of regulation including genomic amplification, transcriptional, posttranscriptional, and posttranslational controls are involved however; the mechanism by which AEG 1 itself induces its oncogenic effects is still poorly understood. Just as questions remain about the exact role of AEG-1 in carcinogenesis, very little is known about the role of AEG-1 in regulating normal physiological functions in the liver. With the help of the Massey Cancer Center Transgenic/Knockout Mouse Core, our lab has successfully created a germline-AEG-1 knockout mouse (AEG-1-/-) as a model to interrogate AEG-1 function in vivo. Here I present the insights gained from efforts to analyze this novel AEG-1-/- mouse model. Aspects of the physiological functions of AEG-1 will be covered in chapter two wherein details of the characterization of the AEG-1-/- mouse are described including the role of AEG-1 in lipid metabolism. Chapter three discusses novel discoveries about the specific role of AEG-1 in mediating hepatocarcinogenesis by modulating NF-κB, a critical inflammatory pathway. First identified over a decade ago, Astrocyte Elevated Gene-1 (AEG-1) has been studied extensively due to early reports of its overexpression in various cancer cell lines. Research groups all over the globe including our own have since identified AEG-1 overexpression in cancers of diverse lineages including cancers of the liver, colon, skin, prostate, breast, lung, esophagus, neurons and neuronal glia as compared to matched normal tissue. A comprehensive and convincing body of data currently points to AEG-1 as an essential component, critical to the progression and perhaps onset of cancer. AEG-1 is a potent activator of multiple pro-tumorigenic signal transduction pathways such as mitogen-activated protein extracellular kinase (MEK)/ extracellular signal-regulated kinase (ERK), phosphotidyl-inositol-3-kinase (PI3K)/Akt/mTOR, NF-κB and Wnt/β-catenin pathway. In addition, studies show that AEG-1 not only alters global gene and protein expression profiles, it also modulates fundamental intracellular processes, such as transcription, translation and RNA interference in cancer cells most likely by functioning as a scaffold protein. The mechanisms by which AEG-1 is overexpressed in cancer have been studied extensively and it is clear that multiple layers of regulation including genomic amplification, transcriptional, posttranscriptional, and posttranslational controls are involved however; the mechanism by which AEG 1 itself induces its oncogenic effects is still poorly understood. Just as questions remain about the exact role of AEG-1 in carcinogenesis, very little is known about the role of AEG-1 in regulating normal physiological functions in the liver. With the help of the Massey Cancer Center Transgenic/Knockout Mouse Core, our lab has successfully created a germline-AEG-1 knockout mouse (AEG-1-/-) as a model to interrogate AEG-1 function in vivo. Here I present the insights gained from efforts to analyze this novel AEG-1-/- mouse model. Aspects of the physiological functions of AEG-1 will be covered in chapter two wherein details of the characterization of the AEG-1-/- mouse are described including the role of AEG-1 in lipid metabolism. Chapter three discusses novel discoveries about the specific role of AEG-1 in mediating hepatocarcinogenesis by modulating NF-κB, a critical inflammatory pathway.

The purpose of this quality improvement project was to increase physical activity among postoperative adult liver transplant patients, improve documentation of daily activity, and ultimately influence quality of life (QOL). Quality of life has been shown to improve dramatically after liver transplant, as patients enter transplant severely debilitated with limitations on their physical activity which carry over following transplant. The literature supports that liver transplant patients should engage in physical activity which may improve QOL. Thirteen liver transplant patients were recruited within the seven days of their post-operative hospitalization. Twelve patients consented, were educated about the benefits of walking, given instructions for how to gradually increase their walking activity, and how to track this activity in a daily log. The International Physical Activity Questionnaire (IPAQ) that calculates level of physical activity (metabolic equivalent or MET score) was conducted at baseline and six weeks. Patients were also asked to rate their perceived quality of life on a ten point scale. Eight patients completed the study with four patients medically unable to complete the walking program. Baseline MET and QOL scores were compared between Time 1 and Time 2 (six weeks). The IPAQ baseline score increased from 407.5 MET to 1,711.5 MET, however, results were not statistically significant. Quality of life improved from Time 1 average score of 5.5 (SD=2.51) to Time 2 average score of 8.25 (SD=1.67) and was statistically significant (P=0.27). Liver transplant patients gradually increased their walking activity over a six week period and documented that activity daily. QOL was also purported to increase which is consistent with findings in the literature. Implementing a post liver transplant walking program is feasible and beneficial for patients and should be a standard of care.

<p>Predictive modeling in healthcare involves the development of data-driven and computational models which can predict what will happen, be it for a single individual or for an entire system. The adoption of predictive models can guide various stakeholders’ decision-making in the healthcare sector, and consequently improve individual outcomes and the cost-effectiveness of care. With the rapid development in healthcare of big data and the Internet of Things technologies, research in healthcare decision-making has grown in both importance and complexity. One of the complexities facing those who would build predictive models is heterogeneity of patient populations, clinical practices, and intervention outcomes, as well as from diverse health systems. There are many sub-domains in healthcare for which predictive modeling is useful such as disease risk modeling, clinical intelligence, pharmacovigilance, precision medicine, hospitalization process optimization, digital health, and preventive care. In my dissertation, I focus on predictive modeling for applications that fit into three broad and important domains of healthcare, namely clinical practice, public health, and healthcare system. In this dissertation, I present three papers that present a collection of predictive modeling studies to address the challenge of modeling heterogeneity in health care. The first paper presents a decision-tree model to address clinicians’ need to decide among various liver cirrhosis diagnosis strategies. The second paper presents a micro-simulation model to assess the impact on cardiovascular disease (CVD) to help decision makers at government agencies develop cost-effective food policies to prevent cardiovascular diseases, a public-health domain application. The third paper compares a set of data-driven prediction models, the best performing of which is paired together with interpretable machine learning to facilitate the coordination of optimization for hospital-discharged patients choosing skilled nursing facilities. This collection of studies addresses important modeling challenges in specific healthcare domains, and also broadly contribute to research in medical decision-making, public health policy and healthcare systems.</p>

Background and aims Heart failure (HF) is a major cause of serious morbidity and death in the population and one of the leading medical causes of hospitalization among people older than 60 years. The aim of this thesis was to describe how to create and how to analyze a Heart Failure Registry with emphasis on Anemia and Quality of Life. (Paper I) We described the creation of the Swedish Heart Failure Registry (SwedeHF) as an instrument, which may help to optimize the handling of HF patients and show how the registry can be used to improve the management of patients with HF. (Paper II) In order to show how to analyze a HF registry we investigated the prevalence of anemia, its predictors, and its association with mortality and morbidity in a large cohort of unselected patients with HFrEF included in the SwedeHF, and to explore if there are subgroups of HF patients identifying high--‐risk patients in need of treatment. (Paper III) In order to show another way of analyzing a HF registry we assessed the prevalence of, associations with, and prognostic impact of anemia in patients with HFmrEF and HFpEF. (Paper IV) Finally we examined the usefulness of EQ--‐ 5D as a measure of patient--‐reported outcomes among HF patients using different analytical models and data from the SwedeHF, and comparing results about HRQoL for patients with HFpEF and HFrEF. Methods An observational study based on the SwedeHF database, consisting of about 70 variables, was undertaken to describe how a registry is created and can be used (Paper I). One comorbidity (anemia) was applied to different types of HF patients, HFrEF (EF &lt;40%) (II) and HFmrEF (EF 40--‐49% ) or HFpEF (&gt; 50%) (III) analyzing the data with different statistical methods. The usefulness of EQ--‐5D as measure of patient--‐ reported outcomes was studied and the results about HRQoL were compared for patients with HFpEF and HFrEF (IV). Results In the first paper (Paper I) we showed how to create a HF registry and presented some characteristics of the patients included, however not adjusted since this was not the purpose of the study. In the second paper (Paper II) we studied anemia in patients with HFrEF and found that the prevalence of anemia in HFrEF were 34 % and the most important independent predictors were higher age, male gender and renal dysfunction. One--‐year survival was 75 % with anemia vs. 81 % without (p&lt;0,001). In the matched cohort after propensity score the hazard ratio associated with anemia was for all--‐cause death 1.34. Anemia was associated with greater risk with lower age, male gender, EF 30--‐39%, and NYHA--‐class I--‐II. In the third paper (Paper III) we studied anemia in other types of HF patients and found that the prevalence in the overall cohort in patients with EF &gt; 40% was 42 %, in HFmrEF 38 % and in HFpEF (45%). Independent associations with anemia were HFpEF, male sex, higher age, worse New York Heart Association class and renal function, systolic blood pressure &lt;100 mmHg, heart rate ≥70 bpm, diabetes, and absence of atrial fibrillation. One--‐year survival with vs. without anemia was 74% vs. 89% in HFmrEF and 71% vs. 84% in HFpEF (p&lt;0.001 for all). Thus very similar results in paper II and III but in different types of HF patients. In the fourth paper (Paper IV) we studied the usefulness of EQ--‐5D in two groups of patients with HF (HFpEF and HFrEF)) and found that the mean EQ--‐5D index showed small reductions in both groups at follow--‐up. The patients in the HFpEF group reported worsening in all five dimensions, while those in the HFrEF group reported worsening in only three. The Paretian classification showed that 24% of the patients in the HFpEF group and 34% of those in the HFrEF group reported overall improvement while 43% and 39% reported overall worsening. Multiple logistic regressions showed that treatment in a cardiology clinic affected outcome in the HFrEF group but not in the HFpEF group (Paper IV). Conclusions The SwedeHF is a valuable tool for improving the management of patients with HF, since it enables participating centers to focus on their own potential for improving diagnoses and medical treatment, through the online reports (Paper I). Anemia is associated with higher age, male gender and renal dysfunction and increased risk of mortality and morbidity (II, III). The influence of anemia on mortality was significantly greater in younger patients in men and in those with more stable HF (Paper II, III). The usefulness of EQ--‐5D is dependent on the analytical method used. While the index showed minor differences between groups, analyses of specific dimensions showed different patterns of change in the two groups of patients (HFpEF and HFrEF). The Paretian classification identified subgroups that improved or worsened, and can therefore help to identify needs for improvement in health services (Paper IV).

Liver disease is now a leading cause of non-AIDS related morbidity and mortality in people living with HIV (PLWH). The present study investigated the interplay between adverse lifestyle factors that are prevalent in PLWH, biological mediators of liver pathogenesis, and a non-invasive measure of liver fibrosis (FIB-4 index) in HIV mono- and HIV/HCV co-infected individuals. The results of this investigation in the Miami Adult Studies of HIV (MASH) cohort show that the odds of liver fibrosis progression significantly increased over two years for HIV mono-infected participants who drank alcohol hazardously (OR 3.038, P=0.048), and had BMI ≥ 28kg/m2 (OR 2.934, P=0.027). Cocaine use reduced the odds of advancing one stage of liver fibrosis (OR 0.228, P=0.038), but an interaction between high BMI and cocaine use slightly raised the odds by 4.8% of liver fibrosis progression (P=0.072). HIV/HCV co-infected participants showed interactions between cocaine use and high BMI with increased FIB-4 stage (OR 4.985, P= 0.034), however no lifestyle factors could independently predict FIB-4 stage in this group. Biological mediators previously associated with liver pathogenesis were associated with higher FIB-4 index over 2 years in a subset of (n=65) HIV mono-infected participants. Plasma measures of oxidative stress (% oxidized glutathione: OR 4.342, P= 0.046), hepatocyte-specific apoptosis (Cytokeratin-18 (CK-18): OR 1.008, P=0.021), and microbial endotoxin (lipopolysaccharide (LPS): OR 1.098, P= 0.097) were associated with having higher odds of progressing at least one stage of FIB-4 over 2 years. The same biological mediators were also associated with liver fibrosis within HIV infected people who also had a harmful lifestyle characteristic. FIB-4 index was significantly associated with % oxidized glutathione in obese subjects (β=0.563, P=0.018), TGF-β1 in cocaine users (β=0.858, P=0.027), and CK-18 in HIV infected individuals without any adverse lifestyle factors (β=0.435, P=0.015). Taken together, the findings of these studies describe interrelationships between HIV disease status, lifestyle, and biological mediators of liver fibrosis. The results show interactions between lifestyle conditions and the mediators of liver fibrosis may account for higher rates of liver disease in HIV infection. Research is warranted to develop personalized therapeutics for PLWH to curb the burden of liver disease.

Kalprotektin är ett protein som bygger upp 60 % av neutrofila granulocyters granula. Vid inflammationer i tarmen vandrar neutrofila granulocyter ut genom mukosan och följer med faeces ut ur kroppen. Proteinet är kalciumbindande och surt och dessa egenskaper utnyttjas i moderna extraktionskit för att mäta mängden kalprotektin i faeces (F-kalprotektin). Thermofisher är ett företag som utvecklat en metod för analys av F-kalprotektin och den kallas för EliA Extraction kit 2 där Kalprotektin extraheras med hjälp av bufferten i röret och analyseras därefter av Phadia 250. Syftet med studien var att utföra en metodjämförelse mellan två metoder. I den första metoden kommer den rutinmetod som används på laboratoriet användas för att upparbeta faeces. I den andra kommer patienterna som skickar sin faeces direkt samlade i extraktionsrören till analys. Syftet är att jämföra analysresultaten för att se om de är lika. Studien utfördes på 26 vuxna patienter varav 11 var kvinnor och 15 var män. Innan analys av prover analyserades tre kontroller för att kontrollera om metoden för analys av F-kalprotektin fick analyseras. Under studien utfördes fyra olika inom-serie precision på fyra olika patienter med 25 replikat för att beräkna variationskoefficienten (CV). Metodens CV varierade mellan 19 - 27 %. Efter att en reparation på instrumentet utförts analyserades två prover med inom-serie precision och CV blev 8 – 13 %. Metodernas resultat hade stark korrelation (R = 0,9055) och vid beräkning av resultat med Mann-Whitney U-test sågs ingen signifikant skillnad (p = 0,3059). Det fanns dock en del prover som hade stor skillnad på resultatet vilket kan ge fel diagnos/klassning. För att kunna bedöma om den nya metoden kan användas som ett alternativ till rutinmetoden måste en större population analyseras. / Calprotectin is a protein that makes up 60 % of the granules of neutrophilic granulocytes. In inflammation of the intestine, neutrophilic granulocytes migrate out through the mucosa and follow the feces out of the body. The protein is calcium-binding and acidic and these properties are used in modern extraction kits to measure the amount of calprotectin in feces  (F-calprotectin). Thermofisher is a company that has developed a method for analysis of F-calprotectin and it is called EliA Extraction kit 2 which is analyzed by Phadia 250. The purpose of the study was to perform a method comparison between two methods. In the first method, the routine method used in the laboratory will be used to process feces. In the second, the patients who send their feces directly collected in the extraction tubes come for analysis. The purpose is to compare the analysis results to see if they are similar. The study was performed on 26 individuals of which 11 were women and 15 were men. Before analyzing, three controls were analyzed to check the method of analysis of F-calprotectin. A negative check and a positive check as well as a calibration check. During the study, four different within-run precision were performed on four different patients with 25 replicates to calculate the coefficient of variation (CV). The CV of the method varied between 19 – 27 %. After a repair on the instrument was performed, the CV was 8 – 13 %. The results of the methods had a strong correlation (R = 0.9055) and when calculating the results with Mann-Whitney U-test, no significant difference was seen (p = 0.3059). In order to be able to assess whether the new method can be used as an alternative to the routine method, a larger population must be analyzed.

Background & Aims: Despite advances in surgical care and chemotherapeutic regimens, the five-year survival rate for Stage IV Hepatoblastoma (HB), the predominant pediatric liver tumor, remains at 27%. YAP1 and β-Catenin co-activation occurs in 80% of children’s HB; however, a lack of conditional genetic models precludes exploration of tumor maintenance and therapeutic targets. Thus, the clinical need for a targeted therapy remains unmet. Given the predominance of YAP1 and β-catenin activation in children’s tumors, I sought to evaluate YAP1 as a therapeutic target in HB. Approach & Results: Herein, I engineered the first conditional murine model of HB using hydrodynamic injection to deliver transposon plasmids encoding inducible YAP1S127A, constitutive β-CateninDelN90, and a luciferase reporter to murine liver. Tumor regression was evaluated using in vivo bioluminescent imaging, and tumor landscape characterized using RNA sequencing, ATAC sequencing and DNA foot-printing. Here I show that YAP1 withdrawal in mice mediates >90% tumor regression with survival for 230+ days. Mechanistically, YAP1 withdrawal promotes apoptosis in a subset of tumor cells and in remaining cells induces a cell fate switch driving therapeutic differentiation of HB tumors into Ki-67 negative “hbHep cells.” hbHep cells have hepatocyte-like morphology and partially restored mature hepatocyte gene expression. YAP1 withdrawal drives formation of hbHeps by modulating liver differentiation transcription factor (TF) occupancy. Indeed, tumor-derived hbHeps, consistent with their reprogrammed transcriptional landscape, regain partial hepatocyte function and can rescue liver damage in mice. Conclusions: YAP1 withdrawal, without modulation of oncogenic β-Catenin, significantly regresses hepatoblastoma, providing the first in vivo data to support YAP1 as a therapeutic target for HB. Modulating YAP1 expression alone is sufficient to drive long-term regression in hepatoblastoma because it promotes cell death in a subset of tumor cells and modulates transcription factor occupancy to reverse the fate of residual tumor cells to mimic functional hepatocytes.