Introdução: O aumento no consumo mundial de imunoglobulina humana tem desafiado os sistemas de saúde no estabelecimento de padrões de utilização adequados a esta terapia. O conhecimento das políticas públicas pelos profissionais de saúde, aderidos a uma conscientização ao uso racional e estudos baseados em evidências científicas é fundamental para assegurar o acesso adequado e uma maior segurança e efetividade de tratamento. Objetivo: Avaliar a utilização de imunoglobulina humana em hospital universitário de alta complexidade do sul do país e suas indicações relacionando as mesmas aos protocolos clínicos estabelecidos. Métodos: Trata-se de um estudo transversal, retrospectivo, baseado na busca de informações através do prontuário eletrônico dos pacientes do Hospital de Clínicas de Porto Alegre no período de janeiro à dezembro de 2015 Resultados: Foram identificadas 191 prescrições de imunoglobulina humana endovenosa, totalizando 116 pacientes. Desses pacientes, 23% apresentaram síndrome de Guillain Barré, púrpura trombocitopênica idiopática, miastenia gravis, transplante renal, imunodeficiência com aumento de IgM e outras anemias hemolíticas autoimunes. Todas essas situações clínicas tem indicação de uso de acordo com os protocolos estabelecidos pelo Ministério da Saúde. Os demais casos identificados (77%) não constam nas indicações previstas nos protocolos do Ministério da Saúde. Conclusão: Foi possível identificar a utilização de imunoglobulina humana endovenosa em hospital de alta complexidade e quantificar os casos clínicos que fazem uso desse medicamento e que apresentam protocolos nacionais orientando os profissionais de saúde quanto a correta administração desse medicamento. Observou-se que a maioria dos casos identificados no estudo não apresentam regulamentos oficiais que autorizem a sua administração. / Introduction: The increase in the world consumption of human immunoglobulin has challenged the health systems in establishing appropriate standards of use for this therapy. Knowledge of public policies by health professionals adhering to rational use awareness and evidence-based studies is critical to ensure adequate access and greater safety and effectiveness of treatment. Objective: To evaluate the use of human immunoglobulin in a university hospital of high complexity in the South of the country and its indications relating them to the established clinical protocols. Methods: This is a cross-sectional, retrospective study based on the search of information through the electronic medical records of patients from the Hospital de Clínicas of Porto Alegre from January to December 2015 Results: 191 prescriptions of intravenous human immunoglobulin were identified, totaling 116 patients. Of these patients, 23% had Guillain Barré syndrome, idiopathic thrombocytopenic purpura, myasthenia gravis, renal transplantation, immunodeficiency with increased IgM and other autoimmune hemolytic anemias. All of these clinical situations are indicated for use according to protocols established by the Ministry of Health. The other cases identified (77%) are not included in the indications provided for in the protocols of the Ministry of Health. Conclusion: It was possible to identify the use of intravenous human immunoglobulin in hospital of high complexity and to quantify the clinical cases that use this medicine and that present national protocols guiding healthcare professionals about the correct administration of this medicine. It was observed that the majority of the cases identified in the study do not present official regulations that authorize its administration.

Introdução: O aumento no consumo mundial de imunoglobulina humana tem desafiado os sistemas de saúde no estabelecimento de padrões de utilização adequados a esta terapia. O conhecimento das políticas públicas pelos profissionais de saúde, aderidos a uma conscientização ao uso racional e estudos baseados em evidências científicas é fundamental para assegurar o acesso adequado e uma maior segurança e efetividade de tratamento. Objetivo: Avaliar a utilização de imunoglobulina humana em hospital universitário de alta complexidade do sul do país e suas indicações relacionando as mesmas aos protocolos clínicos estabelecidos. Métodos: Trata-se de um estudo transversal, retrospectivo, baseado na busca de informações através do prontuário eletrônico dos pacientes do Hospital de Clínicas de Porto Alegre no período de janeiro à dezembro de 2015 Resultados: Foram identificadas 191 prescrições de imunoglobulina humana endovenosa, totalizando 116 pacientes. Desses pacientes, 23% apresentaram síndrome de Guillain Barré, púrpura trombocitopênica idiopática, miastenia gravis, transplante renal, imunodeficiência com aumento de IgM e outras anemias hemolíticas autoimunes. Todas essas situações clínicas tem indicação de uso de acordo com os protocolos estabelecidos pelo Ministério da Saúde. Os demais casos identificados (77%) não constam nas indicações previstas nos protocolos do Ministério da Saúde. Conclusão: Foi possível identificar a utilização de imunoglobulina humana endovenosa em hospital de alta complexidade e quantificar os casos clínicos que fazem uso desse medicamento e que apresentam protocolos nacionais orientando os profissionais de saúde quanto a correta administração desse medicamento. Observou-se que a maioria dos casos identificados no estudo não apresentam regulamentos oficiais que autorizem a sua administração. / Introduction: The increase in the world consumption of human immunoglobulin has challenged the health systems in establishing appropriate standards of use for this therapy. Knowledge of public policies by health professionals adhering to rational use awareness and evidence-based studies is critical to ensure adequate access and greater safety and effectiveness of treatment. Objective: To evaluate the use of human immunoglobulin in a university hospital of high complexity in the South of the country and its indications relating them to the established clinical protocols. Methods: This is a cross-sectional, retrospective study based on the search of information through the electronic medical records of patients from the Hospital de Clínicas of Porto Alegre from January to December 2015 Results: 191 prescriptions of intravenous human immunoglobulin were identified, totaling 116 patients. Of these patients, 23% had Guillain Barré syndrome, idiopathic thrombocytopenic purpura, myasthenia gravis, renal transplantation, immunodeficiency with increased IgM and other autoimmune hemolytic anemias. All of these clinical situations are indicated for use according to protocols established by the Ministry of Health. The other cases identified (77%) are not included in the indications provided for in the protocols of the Ministry of Health. Conclusion: It was possible to identify the use of intravenous human immunoglobulin in hospital of high complexity and to quantify the clinical cases that use this medicine and that present national protocols guiding healthcare professionals about the correct administration of this medicine. It was observed that the majority of the cases identified in the study do not present official regulations that authorize its administration.

Systemic lupus erythematosus (SLE) and Sj?gren?s syndrome (SS) are both heterogeneous autoimmune diseases with strong B cell aspects. A proportion of SLE and SS patients exhibit elevated serum BAFF (B cell activating factor of the TNF family); BAFF plays a key role in B cell homeostasis, survival and tolerance. BAFF transgenic (Tg) mice develop nephritis and salivary gland destruction that resemble aspects of SLE and SS respectively. Autoimmune disease development in BAFF Tg mice correlates with marginal zone (MZ) B cell expansion and the abnormal presence of MZ-like B cells outside of the spleen. The role of MZ B cells in BAFF induced autoimmune disease was analysed by crossing BAFF Tg mice with Lymphotoxin-β knockout mice (creating LTβ-BTg mice) which lack most peripheral lymph nodes, and also lack MZ B cells as a result of disrupted splenic architecture. LTβ-BTg mice were not protected against nephritis but exhibited reduced salivary gland infiltration and destruction. Indicating that the development of sialadenitis but not nephritis in BAFF Tg mice is MZ B cell dependent. Nephritis development in LTβ-BTg mice was associated with the detection of B-1 B cells in the inflamed kidneys. As B-1a B cell survival is dependent on the spleen, the contribution of B-1a B cells to nephritis development in BAFF Tg mice was assessed by crossing BAFF Tg mice to congenitally asplenic Hox11-/- mice (creating Hox11 -BTg mice). The absence of a spleen and B-1a B cells in Hox11-BTg mice delayed the nephritis development. In contrast, splenectomy of BAFF Tg mice at 12 weeks of age did not alter nephritis onset. In these mice B-1a B cells persisted in the peritoneal cavity and MZ-like B cells were detected in the periphery 8 months after surgery. In summary, nephritis development in BAFF Tg mice is unaltered by the absence of MZ B cells, but delayed in the absence of a spleen, MZ and B-1a B cells. Thus, B-1a and B-1b B cells may be potential targets for the treatment of nephritis in SLE patients with elevated BAFF.

Systemic lupus erythematosus (SLE) and Sj?gren?s syndrome (SS) are both heterogeneous autoimmune diseases with strong B cell aspects. A proportion of SLE and SS patients exhibit elevated serum BAFF (B cell activating factor of the TNF family); BAFF plays a key role in B cell homeostasis, survival and tolerance. BAFF transgenic (Tg) mice develop nephritis and salivary gland destruction that resemble aspects of SLE and SS respectively. Autoimmune disease development in BAFF Tg mice correlates with marginal zone (MZ) B cell expansion and the abnormal presence of MZ-like B cells outside of the spleen. The role of MZ B cells in BAFF induced autoimmune disease was analysed by crossing BAFF Tg mice with Lymphotoxin-β knockout mice (creating LTβ-BTg mice) which lack most peripheral lymph nodes, and also lack MZ B cells as a result of disrupted splenic architecture. LTβ-BTg mice were not protected against nephritis but exhibited reduced salivary gland infiltration and destruction. Indicating that the development of sialadenitis but not nephritis in BAFF Tg mice is MZ B cell dependent. Nephritis development in LTβ-BTg mice was associated with the detection of B-1 B cells in the inflamed kidneys. As B-1a B cell survival is dependent on the spleen, the contribution of B-1a B cells to nephritis development in BAFF Tg mice was assessed by crossing BAFF Tg mice to congenitally asplenic Hox11-/- mice (creating Hox11 -BTg mice). The absence of a spleen and B-1a B cells in Hox11-BTg mice delayed the nephritis development. In contrast, splenectomy of BAFF Tg mice at 12 weeks of age did not alter nephritis onset. In these mice B-1a B cells persisted in the peritoneal cavity and MZ-like B cells were detected in the periphery 8 months after surgery. In summary, nephritis development in BAFF Tg mice is unaltered by the absence of MZ B cells, but delayed in the absence of a spleen, MZ and B-1a B cells. Thus, B-1a and B-1b B cells may be potential targets for the treatment of nephritis in SLE patients with elevated BAFF.

Thesis (M.S.)--University of Michigan, 1991.

published_or_final_version / abstract / Pathology / Doctoral / Doctor of Philosophy

BACKGROUND: Patients suffering from inflammatory bowel disease (IBD) are at increased risk of vitamin D deficiency. Daily or weekly vitamin D supplementation has not proven to be effective in improving vitamin D status, and it is thought that this failure has been primarily due to a lack of compliance. Circulating vitamin D is crucial to bone growth and development in children and adolescents. However, more recent data has demonstrated that vitamin D also plays a significant role in the maintenance and regulation of the immune system. OBJECTIVES: The primary aim of this study is to investigate the safety and efficacy of administering high dose oral vitamin D therapy in pediatric patients with IBD. We chose to study patients receiving Remicade, an immunosuppressive monoclonal antibody therapy administered intravenously, as the need for scheduled hospital-based infusions provides a unique opportunity to ensure compliance in our study population. METHODS: We identified consecutive pediatric patients with IBD with a recent 25-hydroxyvitamin D (25OHD) level < 30ng/mL, maintained on Remicade, and with no history of kidney or liver disease for inclusion in the study from November 2017 and November 2018. Enrolled patients received one-year of open-label therapy. Vitamin D treatment doses were assigned by Remicade interval and patients received either 50,000 international units (IU) (every 4-5 weeks) or 100,000 IU (every 6-8 weeks) vitamin D3 orally at the time of their Remicade infusions. In addition to vitamin D levels, spot urine calcium to creatinine ratios, serum calcium, phosphorus, and blood urea nitrogen (BUN) levels, quality of life metrics, and surveys pertaining to dietary vitamin D intake and ultraviolet B (UVB) radiation exposure were collected throughout the study period. RESULTS: Baseline vitamin D status in enrolled patients did not differ by gender, dosing group, diet, or diagnosis (Crohn disease or ulcerative colitis). Subjects reached steady-state serum 25OHD levels after three doses administered over a span of 4 to 8 months, our data demonstrated an increase in average 25OH vitamin D levels from 21.17 ng/mL to 28.19 ng/mL in the 50,000 IU and 23.00 ng/mL to 33.18 ng/mL in the 100,000 IU dose groups, respectively. The improvement in vitamin D status did not correlate with changes in quality of life or disease activity. The response to vitamin D therapy was independent of diet, sun exposure, race, gender, diagnosis, or season of enrollment. There were no adverse events, including changes in urine calcium to creatinine excretion or serum BUN and creatinine values. Several patients manifest a small decrease in serum phosphorus during the initial phase of the study. However, these changes were transient and no subjects exhibited clinical signs or symptoms of hypophosphatemia. CONCLUSION: High dose, interval vitamin D supplementation achieved steady-state 25OHD levels of 30 ng/mL or greater, with no signs of toxicity in patients enrolled in this pilot study. These data suggest that high-dose interval therapy may be a feasible treatment option that bypasses limitations related to difficulties with patient compliance. Further studies are necessary to determine optimal dosage regimens and to assess endpoints related to immune function and improvements to gastrointestinal health.

ANCA-associated vasculitis is a group of devastating autoimmune diseases that predominantly target and destroy small blood vessels. The interaction of neutrophils and monocytes with the endothelial cell lining of blood vessels is imperative to understanding the pathophysiology of the disease. The nature and temporal dynamics of these interactions are mostly unknown and could provide a currently unmet clinical need for more reliable biomarkers of disease activity. This study describes the development of a 4-dimensional in vitro live cell imaging model allowing the interactions of leukocytes with endothelial cells to be analysed in the context of health and disease. Monocytes and neutrophils were isolated from peripheral venous blood of AAV patients and healthy donors. Cells were fluorescently labelled and imaged on a monolayer of human umbilical vein endothelial cells (HUVEC) using a spinning disc confocal microscope. Leukocyte migration, partial and full transmigration, route of transmigration, degranulation and the presence of leukocyte-derived particles inside endothelial cells were measured and the influence of ANCA or BVAS status considered. Following a series of preliminary experiments, it was determined that neutrophil degranulation and partial transmigration indicated early promise as potential biomarkers of disease activity. Several circulating serum analytes correlated with in vitro leukocyte functions, complementing these findings but also highlighting the prevalent immune dysfunction in the pathophysiology and development of the disease. Fatigue is a common symptom within the AAV community and the complex relationship between autoimmune fatigue and leukocyte functions was examined. The data in this thesis describes the development of a novel in vitro live cell imaging platform which can be used to investigate leukocyte functions as potential markers of disease activity as well as understanding their role in the pathophysiology of AAV.

Les maladies auto-immunes sont des maladies chroniques et hétérogènes caractérisées par des réactions du système immunitaire acquis contre les propres tissus sains de l'organisme. Ces maladies affectent presque 5% de la population mondiale et en particulier les jeunes adultes. La complexité de leur spectre est énorme et même si leur étiologie est encore incertaine, il a été démontré que des facteurs génétiques et environnementaux sont impliqués dans le déclenchement du mécanisme pathologique. Cependant, il est nécessaire d'utiliser des outils diagnostiques et / ou pronostiques fiables pour le diagnostic précoce avant que des dommages cellulaires irréversibles ne se produisent et pour surveiller la progression de la maladie.De nombreuses études ont mis en évidence la présence de différents auto-anticorps dans le sérum de patients atteints de maladies auto-immunes. Les auto-anticorps qui sont spécifiques d’une maladie peuvent être utilisés en tant que biomarqueurs (BM) pour son diagnostic alors que les auto-anticorps qui diffèrent en fonction de l'état de la maladie peuvent être utilisés dans le suivi des patients. En fait, dans le cas de l'auto-immunité, un BM facilement détectable et fiable peut être représenté par le titre d'un auto-anticorps spécifique.Dans ce contexte, nous nous intéressons à deux maladies différentes, la sclérose en plaques (SEP) et la gammapathie monoclonale, en utilisant l'approche chimique inverse via le criblage de librairies de peptides par des sérums de patients.En particulier, l'importance des anticorps anti-myéline, et surtout, des anticorps anti-MOG (myéline oligodendrocyte glycoprotéine) est toujours l’objet de débats, soulignant la question très controversée d'un rôle pathogène putatif d'anticorps anti-MOG dans la SEP. Dans cette thèse, nous avons étudié le rôle de MOG comme auto-Ag putatif dans la SEP en utilisant le modèle expérimental d’encéphalomyélite auto-immune (EAE). Ainsi, afin d'évaluer la présence d'un mécanisme d’« epitope spreading » des cellules B, à savoir l'apparition d'une réponse dirigée vers des épitopes distincts de l'agent pathogène induisant la réponse immunitaire, nous avons synthétisé et testé en tant que sondes antigéniques cinq peptides synthétiques qui couvrent la séquence 1-117 de MOG.La seconde étude a porté sur la sélection d'un peptide mimant l'épitope minimal reconnu par l'anticorps monoclonal commercial anti- natural killer cell-1 humain (anti-HNK-1) en utilisant la résonance plasmonique de surface (SPR). L’épitope HNK-1 est considéré comme le déterminant antigénique de la glycoprotéine associée à la myéline (MAG), un composant quantitativement mineur des gaines de myéline. On observe que les patients atteints de troubles neurologiques auto-immuns, tels que la gammapathie monoclonale à IgM et la polyneuropathie démyélinisante, développent souvent des anticorps anti-MAG ciblant spécifiquement l’épitope HNK-1. Par conséquent, l'identification et la caractérisation de ces anticorps est pertinente. Le peptide choisi suite à notre étude pourrait ensuite être utilisé chez des patients atteints de troubles neurologiques pour le développement d'un outil de diagnostic fiable ou de surveillance de l'activité de la maladie par l'identification d'anticorps anti-HNK-1 dans le sérum des patients. / Autoimmune diseases (ADs) refer to chronic and heterogeneous diseases with acquired immune system’s reactions against the body’s own healthy tissues. ADs affect more than 5% of the population worldwide and especially young adults. The complexity of their spectrum is enormous and even if their etiology is still unclear, it was demonstrated that both genetic and environmental factors are involved in triggering the pathological mechanism. Hence, a reliable diagnostic and/or prognostic tool for an early diagnosis of ADs before irreversible cellular damage occurs and for monitoring their progression is demanded.Numerous studies have revealed the presence of different autoantibodies (auto-Abs) in sera of patients suffering from ADs. Autoantibodies that are specific for a disease can be used as biomarkers (BMs) for its diagnosis while autoantibodies that differ depending on the disease state can be used in the follow up of the patients. Actually, in the case of autoimmunity, an easily detectable and reliable BM may be represented by the titer of a specific auto-Ab.In this context, we aimed to identify target(s) of the response for two different ADs, multiple sclerosis (MS) and monoclonal gammopathy, using the chemical reverse approach, which involves the screening of focused antigen (Ag) libraries with patients’ serum.In particular, the significance of anti-myelin antibodies, and especially, anti- Myelin Oligodendrocyte Glycoprotein (anti-MOG) antibodies is still matter of debate, underscoring the highly controversial issue of a putative pathogenetic role of anti-MOG antibodies in MS. In this thesis we investigated the role of MOG as putative auto-Ag in MS using the experimental autoimmune encephalomyelitis (EAE) model. Moreover, in order to assess the presence of a B-cell epitope spreading mechanism, i.e. the occurrence of a response directed toward epitopes distinct from the disease-inducing agent, we synthesized and tested as antigenic probes also five synthetic peptides covering the 1-117 sequence of MOG.The second issue focused on the selection of a peptide mimicking the minimal epitope recognized by the commercial available monoclonal antibody anti-human natural killer cell-1 (anti-HNK-1) using Surface Plasmon Resonance (SPR) technique. HNK-1 epitope, is considered as the antigenic determinant of myelin-associated glycoprotein (MAG), a quantitatively minor component of myelin sheaths. It is observed that patients affected by autoimmune neurological disorders, such as IgM monoclonal gammopathy and demyelinating polyneuropathy, often develop anti-MAG antibodies specifically targeting the HNK-1 epitope. Accordingly, identification and characterisation of these antibodies is relevant. The selected peptide could be subsequently used in earlier stage patients for the development of a novel and reliable diagnostic tool for anti-HNK-1 antibody identification in sera of patients affected by autoimmune neurological disorders monitoring disease activity.

Autoimmune disease is comprised of a wide variety of disorders characterised by a loss of self-tolerance towards a target organ or systemic region leading to its eventual destruction. Type 1 diabetes (T1D), autoimmune thyroid disease (AITD) and inflammatory bowel disease (IBD) are debilitating organ-specific disorders. These disorders arise from a combination of genetic factors and environmental triggers. A greater level of basic understanding of these disorders is required to delay and/or prevent their effects. Numerous autoimmune susceptibility loci have been implicated in the development of these disorders, but only a few causative genes have been identified. The aim of this project was to use comparative mapping between the human and mouse genomes to provide a greater understanding of the human autoimmune susceptibility locus, IDDM6, shown to be involved in a number of autoimmune disease conditions. Hall et al., (2003) previously demonstrated that the mouse autoimmune diabetes locus, Idd21, on distal mouse chr18 contains orthology to human IDDM6, IDDM10, IDDM18 and rat Iddm3. As part of this project the Idd21 locus was fine mapped using the congenic mapping technique. Beginning with the consomic mouse strain, NOD.ABHChr18 (90Mb of Biozzi/ABH-derived diabetes-resistant chr18 introgressed onto a non-obese diabetic (NOD) genetic background), 13 NOD.ABHIdd21 congenic mouse strains were established. The diabetes incidences of these congenic mouse strains were statistically compared stepwise along mouse chr18 and Idd21 was fine mapped to at least four independent autoimmune diabetes loci. Idd21.1 and Idd21.2 were located on distal mouse chr18 in regions orthologous to human IDDM6 and rat Iddm3 and Idd21.3a/b and Idd21.4 were located on proximal mouse chr18 in regions orthologous to human IDDM18 and IDDM10 respectively. Candidate genes of notable interest include Map3k8, Spink5, Cd14, Dcc, Smad4 and 7, Miz1, Nfatc1 and Cd226. Idd21.1 was further fine mapped. Beginning with the NOD.ABHD18Mit8-D18Mit214[(75-85.1Mb)] (Idd21.1) congenic strain (containing at least 10.1Mb of distal chr18 Biozzi/ABH diabetes-resistant DNA introgressed onto a NOD genetic background), seven subcongenic mouse strains were created. The diabetes incidence of these subcongenic mouse strains were statistically compared stepwise along mouse chr18 and Idd21.1 was fine mapped to at least three independent autoimmune diabetes loci; Idd21.11 (72.6-76.1Mb), Idd21.12a/b (75-76.1Mb and 80.6-81.4Mb) and Idd21.13 (84.8-85.1Mb). Candidate genes of interest in these regions include Dcc, Smad4 and 7, Miz1, Nfatc1, and Cd226. Functional characterisation of the Idd21.1 locus was performed by adoptively transferring splenocytes from female NOD or NOD.ABHIdd21.1 mice into cohorts of severe combined immune deficient (scid) female mice, NOD/LtSz.Prkdc[scid] and NOD/LtSz.Prkdc[scid].ABHIdd21.1. There were two notable findings from this work. Firstly, NOD.ABHIdd21.1 splenocytes are not as effective as NOD at transferring diabetes to either NOD/LtSz.Prkdc[scid] (P = 0.0004) or NOD/LtSz.Prkdc[scid].ABHIdd21.1 (P = 0.0178), suggesting that Idd21.1 acquired immune cells are not as diabetogenic as NOD. Secondly, NOD/LtSz.Prkdc[scid].ABHIdd21.1 mice are more resistant to autoimmune attack than NOD/LtSz.Prkdc[scid] when injected with either NOD (P = 0.0015) or NOD.ABHIdd21.1 splenocytes (P = 0.0014), suggesting that Idd21.1 either acts by altering the intrinsic resistance of beta-cells to autoimmune attack or due to changes in the innate immune system. Other NOD-based models of autoimmune disease, spontaneous and experimental autoimmune thyroiditis and spontaneous colitis, were also investigated to determine whether Idd21.1 is a common autoimmune disease locus. When bred onto the NOD.Cg-H2[h4] (thyroiditis model) genetic background Idd21.1 was demonstrated to increase the development of thyroiditis and reduce the incidence of insulitis in spontaneous (untreated) but not experimental (NaI-induced) NOD.Cg-H2[h4] mice. When bred onto the NOD.Cg-Il10[tm1Cgn] (colitis) genetic background Idd21.1 was demonstrated to inhibit the development of rectal prolapse in breeding female NOD.Cg-Il10[tm1Cgn] mice. Data from this thesis demonstrate that the IDDM6 orthologous region in mouse, Idd21.1, contains several loci that influence autoimmune diabetes, thyroiditis and colitis in NOD-based mouse models. These findings are consistent with previous knowledge that IDDM6 is a common autoimmune susceptibility locus.