En France, plus de la moitié des personnes infectées par le VHB ignore leur statut. Le programme " Optiscreen B " visait à évaluer l'intérêt des algorithmes de décision et des tests rapides pour optimiser le dépistage et la prévention de l'hépatite B. Les principaux résultats étaient : La mise en évidence d'occasions manquées de dépistage, principalement liées à une sous-estimation du risque d'exposition lié au pays de naissance.L'application des recommandations de dépistage du CDC pourrait largement diminuer la part des personnes infectées ignorant leur statut mais au prix de très nombreux dépistages.16% des personnes dépistées ont une indication à un traitement antiviral dans les mois suivant le dépistage.100% des personnes ayant des AcHBc isolés avaient une charge virale indétectable.Les TROD AgHBs ont une sensibilité comprise entre 90.5 et 96.5% et une spécificité supérieure à 99%.Le test AcHBs QuickProfileTM a une très bonne spécificité (97.8%) mais une faible sensibilité (58.3%).L'utilité en pratique des TROD VHB pour améliorer la prise en charge des personnes dépistées n'a pas été établie en population générale non ciblée, ceci en partie du fait d'une faible prévalence d'infection et de la sensibilité insuffisante des TROD AcHBs.L'utilisation de TROD AgHBs couplés aux TROD VIH et VHC semble fortement améliorer la cascade de dépistage des personnes migrantes en situation de précarité sociale, à fort risque viral.Les pratiques vaccinales post-dépistage apparaissent extrêmement faibles et le plus souvent liées à l'inertie globale du système.Suite à ce travail, l'analyse médico-économique des différentes stratégies de dépistage est à réaliser. / In France, more than half of those infected with HBV are unaware of their status. The Optiscreen B program aimed to evaluate the interest of testing recommendations and rapid tests (RT) and in optimizing screening, linkage-to-care, and prevention of hepatitis B. The main results were : Identification of missed opportunities for screening, mainly caused by underestimating country of birth as an important risk factor. Implementing screening recommendations from the CDC could significantly reduce the proportion of infected individuals unaware of their status, but with the disadvantage of many tests. 16% of persons testing HBsAg-positive after screening have an indication for antiviral treatment. 100% with isolated anti-HBcAb had undetectable HBV viral load. HBsAg RTs have sensitivities between 90.5 and 96.5% and specificities greater than 99%. The anti-HBsAb QuickProfileTM has very good specificity (97.8%) but low sensitivity (58.3%). The effectiveness of HBV RT in improving linkage-to-care was not established among persons eligible for HBV-screening. This was due in part to low infection prevalence and insufficient sensitivity of the anti-HBsAb RT. Nevertheless, combined use of HBsAg, HIV, and HCV RT seems to greatly improve the cascade of care among migrants in socially precarious situations and at high-risk of infection. HBV-vaccination after HBV screening does not occur often, mainly because of insufficient physician-patient motivation. Increased vaccination coverage might be achieved by emphasizing its need at the organizational level. Following this work, cost-benefit analysis of different screening strategies is greatly needed.

Orientador: Alessandro Lia Mondelli / Resumo: Introdução: O Brasil é classificado pela Organização Mundial de Saúde (OMS) como um país de endemicidade intermediária para o vírus da hepatite C (VHC). Para alcançar melhorias nos serviços de acompanhamento e atendimento pela Equipe Multidisciplinar é necessário cada vez mais o aprimoramento e eficácia nas condutas terapêuticas. Objetivos: Avaliar e comparar o atendimento multidisciplinar de 5 ambulatórios de Hepatite C do Estado de São Paulo; Avaliar o grau de satisfação dos pacientes quanto à qualidade do atendimento recebido por uma equipe multidisciplinar; Descrever o grau de dificuldade no acompanhamento do tratamento dos portadores de hepatite C; Relacionar e comparar a qualidade de atendimento entre os 5 ambulatórios, descrevendo as diferenças entre ambos. Método: estudo transversal, realizado por meio de aplicação de questionário semiestruturado a 425 pacientes dos Ambulatórios, UNESP/HCFMB; UNIFESP/EPM; USP/FMRP; FAMERP/S.J.RIO PRETO e SANTOS CRAIDES. Este trabalho foi aprovado pelo Comitê de Ética em Pesquisa número 1.850.408. Resultados: na avaliação do acompanhamento e atendimento dos serviços ambulatoriais nos diversos centros observaram-se diferenças estatisticamente significantes. A avaliação direta e indireta do atendimento médico e do atendimento de enfermagem foi classificada como boa e ótima, respectivamente. No entanto, os pacientes relataram não sentir diferenças no atendimento de cada profissional. Conclusão: Os resultados desta pesquisa apontam a neces... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Introduction: According with World Health Organization (WHO), Brazil is classified as a country with intermediary endemicity for the hepatitis C virus. In order to improve monitoring and follow-up services by the Multidisciplinary Group it is necessary to improve efficacy on the therapeutic conduct. Objetivos: Evaluate and compare the Multidisciplinary care among ambulatories of Hepatitis C of the State of Sao Paulo, regarding satisfaction monitoring and follow-up services provided by the ambulatories. Methodology: cross-sectional study, conducted by applying a half structuralized questionnaire to 425 patients of the following ambulatories:, CRAIDES/SANTOS; UNIFESP; FAMERP; USPFMRP and HCFMB. Approved by the Ethics in Research Committee. Results: In the evaluation of the follow-up and care of the ambulatorial`s services in the various centers, significant statistical differences were observed. The direct and indirect evaluation of the medical care and nursing care was classified as good and excellent, respectively. However, patients reported no differences in care from one professional to another. Conclusion: The results of this research suggest that, despite the great importance of the theme, in practice, users still demonstrated the lack of knowledge of the multiprofessional team during their treatment. However, they rate the medical and nursing staff during follow-up and outpatient care as good and great. Thus, the results indicate the need for those responsible for outpat... (Complete abstract click electronic access below) / Doutor

ABSTRACT: Hepatitis C virus (HCV) is associated with various glomerulopathies and nephrotic syndrome. However nephrotic syndrome following treatment is rare. Ledipasvir/sofosbuvir (L/S) has recently come into favor in treating HCV due to its relatively mild side effects compared to the more traditional interferon therapy. To the best of our knowledge, there are no reported cases of nephrotic syndrome following treatment with L/S. We present a case of nephrotic syndrome suspected secondary to L/S in a patient with chronic kidney disease. Increased vigilance when assessing therapeutic options in HCV patients with renal comorbidities can improve patient outcomes. A 63 year-old male patient presented to the hospital with shortness of breath, and a two-week history of bilateral lower extremity edema. Past medical history was significant for liver cirrhosis secondary to Hepatitis C genotype Ia, hepatocellular carcinoma status post liver transplantation 6 months prior to admission and Stage 3b chronic kidney disease with baseline creatinine (Cr) approximately 1.5 mg/dl. Medications included L/S for HCV and tacrolimus and prednisone for post-transplant treatment. Patient’s vitals were stable and physical exam was remarkable for facial swelling, mainly on the eyelids, decreased breath sounds bilaterally, distended abdomen with a fluid wave, and 2-3+ pitting edema up to the knees on lower extremities bilaterally. Laboratory work-up was remarkable for low albumin of 3.0 g/dl, and total protein of 5.6 g/ dl. Creatinine of 1.8 mg/dl was elevated from patient’s baseline. HCV viral load was undetectable and electrolytes, transaminases and the complete blood count were within normal limits. Subsequently, urine protein to creatinine ratio was measured because of generalized swelling and hypoproteinemia, which was found to be significantly high at 8.80, compared to 0.04 one year prior. 24-hour total urine protein was found to be 2065 mg/day. Renal ultrasonography showed no hydronephrosis and was otherwise unremarkable. Renal biopsy however, revealed changes suggestive of membranoproliferative glomerulonephritis (MPGN] most likely secondary to HCV. No immune complexes, lambda/kappa light chains, or cryogloblin were appreciated. Nephrotoxic agents such as diuretics and corticosteroids were held. Tacrolimus trough was appropriate to dose level and was continued along with L/S. As admission progressed the patient’s creatinine continued to get worse and rose up to 4.3 mg/dl with persistent proteinuria. With tacrolimus trough levels within normal limits and given L/S was the most recently initiated drug, L/S was thought to be the culprit and was thus held. The renal function began to improve gradually, and the patient was discharged in stable condition with close follow up. Follow up one month later found creatinine and renal function return to baseline and proteinuria resolved. Our case shows that Ledipasvir/sofosbuvir may possibly be related to nephrotic syndrome in HCV patients. Although further studies are needed to prove the causality our case seeks to raise clinical suspicion and increase vigilance when assessing therapeutic options in HCV patients with renal comorbidities such as chronic kidney disease.

abstract: Hepatitis C is an infectious disease that affects 71 million people worldwide and causes liver failure and death if untreated. In 2013, a direct acting antiviral drug, sofosbuvir, revolutionized treatment of the disease. Sofosbuvir showed immense promise, but the high price point at which it was launched created access barriers that prevented it from reaching its full public health potential. By 2016, fewer than 1% of Hepatitis C patients worldwide had received treatment. In the United States (US), concerns about the cost of the drug led public and private payers to implement rationing and treatment restrictions that prevented some of the most vulnerable populations from accessing Hepatitis C treatment at all. Through interviews with researchers, patients and providers, and a literature review of grants, patents, papers, court documents, and news articles, I examine the history of sofosbuvir with attention to the ways in which federal funding practices and intellectual property law encouraged the high initial pricing of the drug. I then examine the impact of this drug on healthcare systems in the United States and abroad, and discuss how the fragmented nature of the United States healthcare system has exacerbated price-based barriers to access. Finally, I discuss intellectual property laws as potential mechanisms to increase access. My study underscores how the political reluctance to use well-established federal funding and intellectual property laws has resulted in a drug development system that delivers medications that are so highly priced that the fragmented US healthcare system cannot compensate for the expense. This leads to low access and poor public health outcomes, and a continued failure to contain or control diseases for which effective therapies exist. / Dissertation/Thesis / Doctoral Dissertation Biology 2020

京都大学 / 新制・課程博士 / 博士(社会健康医学) / 甲第23120号 / 社医博第116号 / 新制||社医||11(附属図書館) / 京都大学大学院医学研究科社会健康医学系専攻 / (主査)教授 今中 雄一, 教授 中山 健夫, 教授 佐藤 俊哉 / 学位規則第4条第1項該当 / Doctor of Public Health / Kyoto University / DFAM

In alcoholic hepatitis (AH), high neutrophil counts correlate with inflammation and poor clinical outcomes. Here, we sought to elucidate the neutrophil-mediated pathogenesis of AH. We revealed that in vivo neutrophil extracellular trap (NET) formation was significantly increased in AH patients and that alcohol alone is sufficient to induce NET formation in neutrophils; thereby, neutrophils increase liver damage through increased NET formation. Moreover, we identify that alcohol-induced NET formation is vital to NETosis and that high-density neutrophils (HDNs) become low-density neutrophils (LDNs) after NET formation in response to alcohol. Through transcriptome profile analysis, we found that genes related to neutrophil activation and immune responses are significantly upregulated in AH HDNs but significantly downregulated in AH LDNs compared to HDNs from healthy subjects. These data suggest that AH HDNs and LDNs have opposing phenotypes: HDNs are activated and more prone to release NETs, while LDNs are functionally incompetent. Consequently, the increase in activated HDNs and defective LDNs are likely associated with an increase in liver damage through NET formation and enhanced susceptibility to infection in AH patients, respectively. Therefore, we evaluated the therapeutic benefits of preventing NET formation in HDNs using peptidyl arginine deiminase 4 (PAD4) inhibition and granulocyte colony-stimulating factor (G-CSF) treatment as well as neutrophil depletion in mice. We observed that in vivo neutrophil depletion and G-CSF treatment prevent NET formation in the liver, thereby significantly reducing liver damage in alcohol-fed mice. Our work identifies the neutrophil/NET-mediated mechanisms of AH pathogenesis and provides insights into therapeutic interventions for AH.

Biological systems, their physical structure and their functions, are built, maintained, and controlled by the activity of enzymes. Understanding how enzymes contribute to the regulation of various pathways and processes allows us to gain a deeper understanding of the entirety of the biological system. As changes in enzyme activity are often essential for the pathogenesis of multiple and varied diseases, identifying these changes represents a crucial step to both understanding the disease and preventing its progression within the individual. Enzymes’ functional output can be controlled by numerous different mechanisms, including control of transcription and translation, subcellular localisation, co-factor interactions, or chemical modification to specific amino acids. Activity-based protein profiling allows the potential for activity of target enzymes to be measured, thereby gaining a more accurate representation of the functional state of the biological system. In this work, profiling differential enzyme activity allows the discovery of previously unknown links between metabolic regulatory enzymes and infection by the hepatitis C virus (HCV). The novel probe wortmannin-yne is described and is shown to be able to report on the activity multiple kinases, including MAPK1, whose activity is dysregulated during HCV replication. Novel probes designed to target a smaller selection of kinases, phosphatidylinositol kinases, are reported and are shown to be capable of measuring HCV-induced changes to not only kinase activity but also regulatory protein-protein interactions with the phosphoinositide kinases. Lastly, the role of microRNA-27b in the HCV-induced dysregulation of lipid metabolic enzymes is examined. Three novel targets of microRNA-27b are identified, and their dysregulation is shown to have an effect on the life cycle of HCV. Altogether, this work has developed new tools for the study of metabolic enzymes and identified new avenues of investigation into the dysregulation of lipid metabolism.

Hepatitis C Virus (HCV) is an infection that can have grave consequences when left untreated. Hepatitis C can be easily eradicated with direct acting antiviral therapy. People who inject drugs (PWID) and inmates are among those with the highest incidence of HCV. However, cure rates among this population remains low. This is, in part, related to an interruption in the HCV care cascade such that only 30% of PWID are linked to care and only 8% of those receive treatment. Inadequate screening and failure to be linked to HCV care remain the largest impediments to treatment success. There is limited research on barriers and facilitators to primary care, where screening may take place, and linkage to HCV care among PWID. Few studies have evaluated vulnerable populations such as those living in rural communities or inmates. The purpose of this dissertation was to develop a broader understanding of barriers and facilitators to healthcare utilization among PWID at the primary care and specialist levels (linkage to care). Three manuscripts addressed important gaps in knowledge. The first was a review of the literature to describe the state of science on linkage to care among PWID. All but one reviewed study recruited from countries with universal healthcare, urban areas, and opioid substitution facilities. The review of the literature revealed that little is known about the barriers/facilitators to linkage to HCV care faced by rural-dwelling PWID from countries without universal healthcare. The second manuscript is a study to determine whether predictors of linkage to care identified in urban-dwelling PWID from countries with universal healthcare predicted seeking HCV care among PWID living in rural Appalachia. Data were obtained from a subsample of 63 HCV positive PWID who recently used opioids, were between the ages of 18-35 years, and lived in one of five rural counties in Kentucky. Logistic regression revealed that recent injection drug use was the only predictor of seeking HCV care. However, remote use of opioid substitution therapy and no transportation issues approached clinical significance. Although not evaluated in our second manuscript, seeing a primary care provider (PCP) is associated with an increased likelihood of being linked to care and higher rates of screening/diagnosis. Among rural dwelling PWID, there are subpopulations that may face unique barriers to linkage to care. One sub-population that may be particularly vulnerable are female PWID who are incarcerated. Therefore, the purpose of the third study was to determine predictors of primary care use using data from 302 female inmates from rural Appalachia with a history of injecting drugs. Age, insurance issues, and health problems that interfere with responsibilities were predictors of PCP use. In this dissertation, I have addressed important gaps in the literature by determining barriers and facilitators to seeking HCV care and primary care use among PWID from rural Kentucky. Additional studies are needed using a larger sample of rural PWID to confirm our findings. In addition, further studies should evaluate system and provider level barriers to linkage to care and PCP use among rural PWID.

Le virus de l’hépatite B (VHB) utilise la machinerie transcriptionnelle nucléaire pour sa réplication. Le génome viral est transporté de la périphérie cellulaire à l’enveloppe nucléaire. Généralement, ce transport intracytoplasmique rétrograde est facilité par le réseau de Mt via l’utilisation du complexe moteur de la dynéine. Nous avons montré que le transport des capsides du VHB dépend des Mt, ce qui permet l’adressage des capsides aux complexes du pore nucléaire (NPC) ; lequel est requis pour l’étape de libération du génome de la capside dans le noyau.Dans cette étude, nous avons utilisé des capsides provenant de virus récupérés dans du surnageant de HepG2.2.15, qui contiennent le génome mature partiellement double brin (capsides matures), et des capsides exprimées chez E.coli. Ces dernières sont utilisées telles quelles, capsides E.coli contenant de l’ARN, ou bien sont utilisées pour préparer des capsides vides. Après microinjection dans des ovocytes de Xenopus laevis, nous avons observé que les capsides vides et les capsides matures sont transloquées aux NPC avec une cinétique similaire. Les capsides contenant de l’ARN ne sont pas identifiées aux NPCs ce qui implique que le transport des deux autres types de capsides est actif. Cela a été confirmé par la pré-injection d’anticorps anti tubuline qui neutralisent le transport assuré par les Mt.L’attachement spécifique des capsides matures et vides aux Mt a été confirmé en utilisant des Mt polymérisés in vitro, nous avons montré que cette interaction nécessitait des protéines cytosoliques. En utilisant des expériences de coïmmunoprécipitation et de cosédimentation nous avons identifié une chaîne légère de la dynéine (DynLL1 membre de la famille Lc8) comme partenaire des capsides. Dans les expériences de microinjection, la comicroinjection d’un excès de DynLL1 avec les capsides inhibe leur transport vers les NPCs, indiquant que DynLL1 est impliquée dans le transport actif des capsides.DynLL2 qui n’interagit pas avec les capsides diffère de DynLL1 de seulement six acides aminés. Par mutagénèse dirigée de DynLL1, nous avons montré l’implication de deux acides aminés dans l’interaction directe avec les capsides. Ces deux acides aminés sont présents à la surface du dimère de DynLL1 et absents dans le sillon résultant de la dimérisation de DynLL1, sillon impliqué dans l’interaction avec la DynIC. Nous avons partiellement reconstitué le complexe DynIC, DynLL1 et capsides vides qui doit en partie refléter la situation in vivo. / Hepatitis B virus (HBV) needs the nuclear transcription machinery for replication. The virus thus depends on the transport of its genome from the cell periphery to the nuclear envelope. In general this retrograde intracytoplasmic trafficking is facilitated along Mt (MT) using motor protein complexes of the dynein family. As we showed earlier HBV capsid transport also depends upon intact MT in order to allow their arrival at the nuclear pores, which in turn is required for genome liberation from the capsid.In the analysis we used virus-derived HBV capsids obtained from the supernatant of HepG2.2.15, which contain the mature partially double-stranded DNA genome (mature capsids) and capsids expressed in E. coli. The latter were applied in two forms: as unspecific E. coli RNA- containing capsids and as empty capsids. Upon microinjection into Xenopus laevis oocytes we observed that mature and empty capsids were translocated to the nuclear pores with a similar kinetic. RNA-containing capsids failed to arrive at the pores implying that transport of the two other capsid types was active. Active translocation was confirmed by pre-injecting anti tubulin antibodies which interfere with MT-mediated translocation.In vitro reconstitution assays confirmed the specific attachment of mature and empty capsids to MTs and showed the need of further cytosolic proteins. Using pull-down and co-sedimentation experiments we identified one dynein light chain (DYNLL1, member of the Lc8 family) as interaction partner of the capsids. Injecting an excess of recombinant DYNLL1 with empty capsids into Xenopus laevis oocytes inhibited capsid transport to the nuclear pores indicating that DYNLL1 was only functional interaction partner implied in active transport.DNYLL2 did not interact with the capsids although differing from DYNLL1 by just six amino acids. Site directed mutagenesis of DYNLL1 revealed that two amino acids were critical for a direct interaction with the capsids. Both localized at the exterior of the DYNLL1 dimer and not in the groove of DYNLL1, which interacts with the dynein intermediate chain. Accordingly we could reconstitute a complex consisting of empty capsids, DYNLL1 and dynein intermediate chain as it should be in the in vivo situation.

Determina la relación entre el nivel de conocimientos sobre hepatitis B y el estado de vacunación de los estudiantes de medicina que formarán parte del programa internado médico 2019 del Hospital Nacional Dos de Mayo (HNDM). Realiza un estudio transversal, analítico, observacional y correlacional. La población está constituida por los 120 estudiantes de medicina que formarán parte del programa internado médico del 2019 del HNDM. Del total de estudiantes de medicina el 61% presentan un mal conocimiento sobre el virus de la hepatitis B (VHB); el 53% presentan estado de vacunación incompleto. Asimismo, se aprecia que del total de estudiantes que recibieron vacunación el 86,2% tienen edad de 22 a 27 años; el 66% son de sexo femenino; el 79.8% provienen de universidad privada; el 71.3% recibió tres dosis de la vacuna contra el VHB y el 88.3% no presentaron eventos supuestamente atribuidos a la vacunación e inmunización (ESAVI). De los estudiantes que tienen un mal conocimiento sobre el VHB el 54,1% tiene un esquema de vacunación incompleta, mientras que el 76% de los estudiantes con conocimiento regular poseen un esquema incompleto. Finalmente, del total de estudiantes con un buen conocimiento sobre el VHB el 92,9% presenta un esquema de vacunación completa. Concluye que el nivel de conocimientos sobre el VHB está relacionado con el estado de vacunación del estudiante de medicina ingresante al programa de internado médico 2019 del HNDM. / Tesis