Microvascular issues can precede major macrovascular diseases and their measurements can predict long-term cardiovascular disease and survival. Therefore, early interventions on microvascular... Show moreMicrovascular issues can precede major macrovascular diseases and their measurements can predict long-term cardiovascular disease and survival. Therefore, early interventions on microvascular issues can potentially delay later cardiovascular problems. Microcirculation is critical for tissue perfusion, exchange of nutrients, oxygen and carbon dioxide between blood and interstitial fluid, and in vascular homeostasis. Assessing the microcirculation is challenging due to spatially heterogeneous structure and variability in perfusion over time and under different conditions.The thesis aims to evaluate standardized techniques for investigating microcirculation. Although numerous techniques are used to assess microvasculature, there's no consensus on which is the most suitable for evaluating microcirculation. The dissertation also explores the possibilities and impossibilities of measuring the microvasculature for pharmacodynamic outcomes of new interventions and medications, and highlights the need for further research to develop effective interventions targeting microcirculation. Show less
Patients with sickle cell disease (SCD) experience a considerable physical and psychosocial disease burden. In recent years, the application of allogeneic hematopoietic stem cell transplantation ... Show morePatients with sickle cell disease (SCD) experience a considerable physical and psychosocial disease burden. In recent years, the application of allogeneic hematopoietic stem cell transplantation (HSCT) to treat adults with SCD has increased. A thorough understanding of patients' physical, mental, and social health before and after cure is needed to meet the needs of this growing group of patients. We aimed to explore the perspectives of adult SCD patients on the changes in their experienced health and personal life goals after being cured. A mixed-methods approach was used, comprising a semistructured interview and a set of 9 Patient Reported Outcomes Measurement Information System (PROMIS) measures. Adult SCD patients who underwent HSCT at least 1 year earlier were eligible to participate in the study. Interviews were thematically analyzed using MAXQDA software. PROMIS T scores were compared with reference scores of the general population using SPSS Statistics. Ten patients participated in the study; their median age was 29.5 years (range, 19 to 49 years), and their median time since HSCT was 2.7 years (range, 1.0 to 3.5 years). Themes from the interviews were (1) pain/living pain free, (2) physical wellbeing, (3) mental well-being, (4) perspective/outlook, (5) education/work, (6) family/friends, and (7) activities/participation. Following the PROMIS framework, we described these themes in a narrative synthesis according to health domain and categorized in 4 chronological time phases: before HSCT, first year post-transplantation, current situation, and future expectations. Physical health improved greatly, but transplantation-related toxicity, ongoing pain from avascular osteonecrosis, and fatigue negatively impacted quality of life in some patients. Furthermore, emotional struggles during the post-transplantation period were common, and patients expressed a need for psychological help. Patients reported improvements in social health and the ability to pursue personal life goals. The mean T scores of all PROMIS measures fell within the normal symptom limits compared with reference data of the general population, although, large variations were observed among the participants, matching our qualitative findings. In general, adult SCD patients experienced improved physical, mental, and social health after cure by HSCT and were able to pursue personal life goals. Yet they found themselves confronted with a new and unfamiliar reality that brought different challenges. Pain due to irreversible avascular osteonecrosis continued to have a negative impact. Clinicians should aim to help patients have realistic expectations before transplantation and offer timely psychological care. (c) 2023 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. Show less
Cruz, L.J.; Dijk, T. van; Vepris, O.; Li, T.M.W.Y.; Schomann, T.; Baldazzi, F.; ... ; Eich, C. 2021
Ex vivo gene editing of CD34+ hematopoietic stem and progenitor cells (HSPCs) offers great opportunities to develop new treatments for a number of malignant and non-malignant diseases. Efficient... Show moreEx vivo gene editing of CD34+ hematopoietic stem and progenitor cells (HSPCs) offers great opportunities to develop new treatments for a number of malignant and non-malignant diseases. Efficient gene-editing in HSPCs has been achieved using electroporation and/or viral transduction to deliver the CRISPR-complex, but cellular toxicity is a drawback of currently used methods. Nanoparticle (NP)-based gene-editing strategies can further enhance the gene-editing potential of HSPCs and provide a delivery system for in vivo application. Here, we developed CRISPR/Cas9-PLGA-NPs efficiently encapsulating Cas9 protein, single gRNA and a fluorescent probe. The initial 'burst' of Cas9 and gRNA release was followed by a sustained release pattern. CRISPR/Cas9-PLGA-NPs were taken up and processed by human HSPCs, without inducing cellular cytotoxicity. Upon escape from the lysosomal compartment, CRISPR/Cas9-PLGA-NPs-mediated gene editing of the gamma-globin gene locus resulted in elevated expression of fetal hemoglobin (HbF) in primary erythroid cells. The development of CRISPR/Cas9PLGA-NPs provides an attractive tool for the delivery of the CRISPR components to target HSPCs, and could provide the basis for in vivo treatment of hemoglobinopathies and other genetic diseases. Show less
Tozatto-Maio, K.; Torres, M.A.; Degaide, N.H.S.; Cardoso, J.F.; Volt, F.; Pinto, A.C.S.; ... ; Gluckman, E. 2020
Sickle cell disease (SCD) is the most common inherited hemoglobinopathy. Hematopoietic stem cell transplantation (HCT) is the sole curative therapy for SCD, but few patients will have a matched... Show moreSickle cell disease (SCD) is the most common inherited hemoglobinopathy. Hematopoietic stem cell transplantation (HCT) is the sole curative therapy for SCD, but few patients will have a matched sibling donor. Patients with SCD are mostly of African origin and thus are less likely to find a matched unrelated donor in international registries. Using HaploStats, we estimated HLA haplotypes for 185 patients with SCD (116 from a Brazilian center and 69 from European Society for Blood and Marrow Transplantation [EBMT] centers) and classified the ethnic origin of haplotypes. Then we assessed the probability of finding an HLA-matched unrelated adult donor (MUD), considering loci A, B, and DRB1 (6/6), in international registries. Most haplotypes were African, but Brazilians showed a greater ethnic admixture than EBMT patients. Nevertheless, the chance of finding at least one 6/6 potential allelic donor was 47% for both groups. Most potential allelic donors were from the US National Marrow Donor Program registry and from the Brazilian REDOME donor registry. Although the probability of finding a donor is higher than previously reported, strategies are needed to improve ethnic diversity in registries. Moreover, predicting the likelihood of having an MUD might influence SCD management. (C) 2020 American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. Show less
Hemoglobinopathies (HBP) are the most common autosomal recessive genetic disorder in Oman. Carriers are usually asymptomatic but carrier couples are at 25% risk of getting a severely affected child... Show moreHemoglobinopathies (HBP) are the most common autosomal recessive genetic disorder in Oman. Carriers are usually asymptomatic but carrier couples are at 25% risk of getting a severely affected child. Public health authorities have focused not only on state of the art management and patient care but also on prevention. The focus of this thesis is to study the molecular spectrum of HBP and the associated genetic determinants to work towards the development of prevention strategies for severe HBP__s in Oman. We have defined the molecular spectrum of the disease all around the country, including beta, alpha and delta globin gene mutations. Furthermore, genotype/phenotype correlation studies were investigated in patients with sickle cell disease (SCD) by looking at determinants such as haplotype/sub-haplotype, alpha thalassemia and hydroxyurea response based on XmnI polymorphism. Identifying genetic determinants is necessary for prognostic purposes, accurate diagnosis and planning for the best tailored treatment to the affected patients. While providing tools for a better care and a better insight on the management of these severe diseases in Oman, the results from this thesis will help to facilitate the prevention of HBP in the country. Show less
Hemoglobinopathies (HbP) are recessive hereditary disorders of hemoglobin, characterized by microcytic hypochromic anemia. HbP diagnostics encompasses three specialties: hematological, biochemical... Show moreHemoglobinopathies (HbP) are recessive hereditary disorders of hemoglobin, characterized by microcytic hypochromic anemia. HbP diagnostics encompasses three specialties: hematological, biochemical and molecular testing. Results of all tests together form the complete diagnosis. The main objective of this thesis was to improve post- and prenatal diagnostics of the hemoglobinopathies. Several molecular assays have been designed, tested and validated. In addition, a number of informative hemoglobinopathy cases have been studied in detail. Diagnostics for hemoglobinopathies is strongly improved over the recent years. In particular the implementation of the MLPA technique made it possible to detect deletions and duplications in the globin gene clusters in patients who remain undiagnosed by applying the conventional techniques. The aCGH technology was developed in order to characterize the breakpoints of novel deletions detected by MLPA more precisely. This has led to the design of several relatively simple gap-PCR assays, which are useful in laboratories where MLPA and aCGH is not available. In addition, gap-PCR can be used for quick screening for the more locally occurring deletions or in family studies. A non-invasive prenatal diagnosis assay for hemoglobinopathies was developed by combining the PAP and MCA techniques. This method will be implemented in the current flow for prenatal diagnosis and will eventually make 50% of the invasive procedures redundant. Show less
Over the last two decades, there has been substantial progress in the area of blood safety in Uganda. In contrast, little attention has been paid to transfusion safety in Uganda and there are gaps... Show moreOver the last two decades, there has been substantial progress in the area of blood safety in Uganda. In contrast, little attention has been paid to transfusion safety in Uganda and there are gaps in laboratory and clinical transfusion practices within hospitals. Assessment of the current practice at Mulago and Mbarara Referral Hospitals showed inadequate documentation of the transfusion process, and poor monitoring of blood recipients. Our findings also indicated that one in every 16 transfused Ugandans and a similar number of RhD negative pregnant women possessed clinically significant red blood cell (RBC) alloantibodies in their plasma. However, RBC alloantibody screening is not performed during pre-transfusion and antenatal testing in the country. Thus, alloimmunized recipients and babies of RhD negative mothers are at high risk of morbidity and mortality due to haemolytic transfusion reactions (HTRs) and haemolytic disease of the fetus and the newborn (HDFN). Furthermore, data on the occurrence of acute and delayed HTRs and HDFN in Uganda are lacking. A cost-effectiveness analysis showed that introduction of RBC alloantibody screening would be cost-effective and improve blood transfusion safety. Therefore, there is need to improve immunohaematological testing in Uganda so that RBC alloimmunization and the consequences thereof may be prevented. Show less
