Efeito de polimorfismos do gene da interleucina-1 (IL-1), IL-6, IL-10 e fator de necrose tumoral na ocorrencia de reação transfusional febril não hemolitica

Orientador: Sara Teresinha Olalla SaadTese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias MedicasResumo: Citocinas inflamatórias como IL-1a e ß, IL-1RA, IL-6, IL-10 e TNF estão envolvidas na resposta imune inclusive na ocorrência de reação febril não hemolítica. Podem acumular nos concentrados de plaquetas (CPs) durante o armazenamento ou serem liberadas por células do receptor de transfusão. Alguns polimorfismos dos genes destas citocinas interferem na produção frente a estímulos. O presente trabalho tem como objetivo avaliar a importância dos polimorfismos IL1B-511C/T, IL1+3953C/T, IL1RN intron2 VNTR, IL6-174G/C, IL10-1082G/A, IL10-819C/T, TNF308G/A e TNF+252G/A em pacientes multitransfundidos e em doadores de sangue, utilizado para produção de CPs, através da avaliação da ocorrência precoce de RFNH associada a transfusão de concentrado de hemácias ou do acúmulo de IL-1a e ß, IL-1RA e TNF nos CPs. Utilizamos o método PCR e digestão com enzimas de restrição (PCR-RFLP) ou sequenciamento automatizado para identificações destes polimorfismos. Em nossa amostra de população da região de Campinas, São Paulo, encontramos freqüências genotípicas comparáveis às populações européia e norte americana e diferenças significativas das populações asiática e de origem africana. O raro alelo IL1RN*0 foi identificado em nosso grupo, não foi demonstrado interferência deste alelo, em heterozigose, na expressão do gene IL1RN em modelo in vitro. Nos CPs obtidos de plasma rico em plaquetas (CP-PRP) foi observado maior acúmulo de IL-1ß nos doadores IL1B-511CC, evidenciado no D5 e D7 de armazenamento. Foi observada correlação inversa entre o conteúdo de plaquetas nos CP-PRP e o acúmulo de IL-1ß e TNF, não havendo correlação com a contaminação leucocitária. Não houve acúmulo de IL-1a e IL- 1RA durante o armazenamento. No grupo de pacientes multitransfundidos, o alelo IL1RN*2 e o genótipo IL1RN*2.2 correlacionaram-se com a ocorrência de RFNH precoce. Concluímos que, o estudo de características genéticas dos doadores de sangue e pacientes pode ajudar a esclarecer as variações de ocorrência e gravidade dos efeitos adversos associados à transfusão de sangue, identificando grupos de maior risco e delineia uma nova linha de investigaçãoAbstract: Inflammatory cytokines such as IL-1a e ß, IL-1RA, IL-6, IL-10 and TNF have been related with immune responses including febrile non-hemolytic reactions. These biological mediators may accumulate in platelet concentrates (PCs) during storage or be secreted by receptor immune cells. Many cytokine gene polymorphisms interfere in the production of cytokines after stimulus. The present study aims to value the significance of polymorphisms IL1B-511C/T, IL1B+3953C/T, IL1RN intron2 VNTR, IL6-174G/C, IL10-1082G/A, IL10-819C/T, TNF308G/A and TNF+252G/A in multitransfused patients and in PCs donors using the evaluation of precocious FNHTR occurrence associated with red blood cells (RBCs) transfusion or accumulations of IL-1a e ß, IL-1RA and TNF in PCs during storage. The methods used were based on PCR and restriction enzymes (PCR-RFLP) or automated DNA sequencing for polymorphism characterizations. In our population sample from Campinas, southeast Brazilian region, we detected similar genotype frequencies to European and North American populations and significant differences to the Asiatic and African ones. A rare allele IL1RN*0 was detected in our control group, we did not demonstrate the interference of this allele in heterozygousity, in the IL1RN gene expression in the in vitro model. In PCs from plasma rich in platelet (PRP-PC), we observed increased IL-1ß accumulation in PCs from IL1B-511CC blood donors on storage days 5 and 7 (D5 and D7). IL-1ß and TNF levels were inversely correlated with platelet content in PRP-PCs and no correlation with leukocyte content. There was no increase in IL-1a e IL-1RA levels during storage. In the multitransfused patients group, the IL1RN*2 allele and IL1RN*2.2 genotype were correlated with the occurrence of precocious FNHTR. We concluded that studies of genotypic characteristics of blood donors and patients might contribute to explain the variations in frequencies and gravity of side effects related with blood transfusion characterizing high risk groups and may outline a new and promising investigation fieldDoutoradoHematologia e HemoterapiaDoutor em Clínica Médic

Prevalence of transfusion-transmitted Chagas disease among multitransfused patients in Brazil

<p>Abstract</p> <p>Background</p> <p>Blood transfusion has always been an important route for Chagas Disease (CD) transmission. The high prevalence of CD in Latin America and its lifelong asymptomatic clinical picture pose a threat for the safety of the blood supply. The outcome of measures designed to improve transfusion safety can be assessed by evaluating the prevalence of CD among multitransfused patients</p> <p>Methods</p> <p>In order to assess the impact of CD control measures on the safety of the blood supply, an observational cross-sectional study was designed to determine the prevalence of CD in 351 highly transfused patients, in which vectorial transmission was excluded. This study compared patients that received transfusion products before (n = 230) and after (n = 121) 1997, when measures to control transfusion-transmitted CD were fully implemented in Brazil.</p> <p>Results</p> <p>The study group consisted of 351 patients exposed to high numbers of blood products during their lifetime (median number of units transfused = 51, range 10–2086). A higher prevalence of transfusion-transmitted CD (1.30%) was observed among multitransfused patients that received their first transfusion before 1997, compared with no cases of transfusion-transmitted CD among multitransfused patients transfused after that year. The magnitude of the exposure to blood products was similar among both groups (mean number of units transfused per year of exposure = 25.00 ± 26.46 and 23.99 ± 30.58 respectively; P = 0.75, Mann-Whitney test).</p> <p>Conclusion</p> <p>Multiple initiatives aimed to control vector and parental transmission of CD can significantly decrease transfusion-transmitted CD in Brazil. Our data suggest that mandatory donor screening for CD represents the most important measure to interrupt transmission of CD by blood transfusions.</p

Cytokine polymorphisms in sickle cell disease and the relationship with cytokine expression

Sickle cell disease is a chronic inflammatory condition characterized by elevated levels of inflammatory cytokines, which may be regulated by genetic polymorphisms and could be associated with diverse disease presentations and alloimmunization. The aim of this study was to evaluate Treg and Th17 cell frequencies, cytokine gene polymorphisms, and their association with cytokine expression profile in patients with sickle cell disease. For that purpose, we evaluated the IL intron 3 variable number tandem repeat (VNTR, genotypes 1.1, 1.2, 2.2, and 2.3), IL4-T590C>T, IL6-174G>C, TNF alpha-308G>A, IL10-819T>C, IL10-592A>C, and IL10-1082A>G polymorphisms and their correlation with TGF beta, IL4, IL6, and IL10 gene expression in sickle cell patients. We observed a significant decrease in Treg frequency together with a substantial increase in Th17 response in patients with sickle cell disease compared with healthy controls (p < 0.001 and p = 0.014, respectively). There was also a higher prevalence of the IL4-590T/T genotype in patients with sickle cell disease than in Afro--Brazilian descendent controls (p < 0.001) and higher expression of IL4 in patients with the 1.1 genotype of IL4 intron 3 VNTR (p = 0.06). Significantly greater gene expression of TGF beta, IL6, and IL10 was observed in sickle cell patients when compared with controls (p = 0.01, 0.03, and <0.001, respectively). Moreover, higher levels of interleukin-6 and -10 were observed in the group of alloimmunized patients. These new data bring insights into the deregulation in the immune system affecting sickle cell patients and must be further investigated in larger cohorts to better characterize individual variations in immune responses and new markers for disease morbidity. Copyright (C) 2016 ISEH - International Society for Experimental Hematology447583589sem informaçã

Low SARS‐CoV‐2 seroprevalence in a cohort of Brazilian sickle cell disease patients: Possible effects of emphasis on social isolation for a population initially considered to be at very high risk

Abstract Despite being initially considered at higher risk for severe COVID‐19, sickle cell disease (SCD) patients have mostly presented clinical severity similar to the general population. As their vulnerability to become infected remains uncertain, we assessed the seroreactivity for SARS‐CoV‐2 to estimate the prevalence of infection and possible phenotypic and socioeconomic determinants for their contagion. Serologic evaluation was performed on 135 patients with an overall prevalence of 11%; positivity was associated with older age and use of public transportation. We speculate that social distancing instructions recommended by our clinic may have contributed to lower levels of infection, but potential protection factors need further investigation

Evaluation of Molecular Methods to Identify Chagas Disease and Leishmaniasis in Blood Donation Candidates in Two Brazilian Centers

In Brazil, blood donation is regulated by the Brazilian Ministry of Health, and all States follow the same protocol for clinical and laboratory screening. Brazil is an endemic country for Chagas disease (CD), caused by Trypanosoma cruzi, and for leishmaniasis, caused by a species of Leishmania spp. Screening for leishmaniosis is not routinely performed by blood banks. Given the antigenic similarity between T. cruzi and Leishmania spp., cross-reactions in serological tests can occur, and inconclusive results for CD have been found. The objective of this study was to apply molecular techniques, e.g., nPCR, PCR, and qPCR, to clarify cases of blood donation candidates with non-negative serology for CD and to analyze the difference between the melting temperature during real-time PCR using SYBR Green. Thirty-seven cases that showed non-negative results for CD using chemiluminescent microparticle immunoassay (CMIA) tests from blood banks in Campo Grande, MS, and Campinas, SP, were analyzed. In the serum samples, 35 samples were evaluated by ELISA, and 24.3% (9/35) showed positive results for CD. nPCR was able to detect 12 positive results in 35 samples (34.28%). qPCR for T. cruzi was quantifiable in the samples that showed a value ≥0.002 par eq/mL (parasite equivalents per milliliter), and in 35 samples, 11 (31.42%) were positive. Of all evaluated samples using the described tests (CMIA, ELISA, nPCR, and qPCR), 18 (48.6%) were positive for CD. For MCA by qPCR, the melting temperature was 82.06 °C ± 0.46 for T. cruzi and 81.9 °C ± 0.24 for Leishmania infantum. The Mann–Whitney test showed a significant value of p T. cruzi and L. infantum could not be considered due to temperature overlap. For leishmaniasis, of the 35 samples with non-negative serology for CD tested by the indirect fluorescent antibody test (IFAT), only one sample (2.85%) was positive (1:80). The PCR for Leishmania spp. was performed on 36 blood samples from donation candidates, and all were negative. qPCR for L. infantum showed 37 negative results for the 37 analyzed samples. The data presented here show the importance of performing two different tests in CD screening at blood banks. Molecular tests should be used for confirmation, thereby improving the blood donation system

Clearance of Persistent SARS-CoV-2 RNA Detection in a NFκB-Deficient Patient in Association with the Ingestion of Human Breast Milk: A Case Report

Currently, there are no evidence-based treatment options for long COVID-19, and it is known that SARS-CoV-2 can persist in part of the infected patients, especially those with immunosuppression. Since there is a robust secretion of SARS-CoV-2-specific highly-neutralizing IgA antibodies in breast milk, and because this immunoglobulin plays an essential role against respiratory virus infection in mucosa cells, being, in addition, more potent in neutralizing SARS-CoV-2 than IgG, here we report the clinical course of an NFκB-deficient patient chronically infected with the SARS-CoV-2 Gamma variant, who, after a non-full effective treatment with plasma infusion, received breast milk from a vaccinated mother by oral route as treatment for COVID-19. After such treatment, the symptoms improved, and the patient was systematically tested negative for SARS-CoV-2. Thus, we hypothesize that IgA and IgG secreted antibodies present in breast milk could be useful to treat persistent SARS-CoV-2 infection in immunodeficient patients

International Society of Blood Transfusion survey of experiences of blood banks and transfusion services during the COVID-19 pandemic

Background and Objectives: The coronavirus disease 2019 (COVID-19) pandemic has impacted blood systems worldwide. Challenges included maintaining blood supplies and initiating the collection and use of COVID-19 convalescent plasma (CCP). Sharing information on the challenges can help improve blood collection and utilization. Materials and Methods: A survey questionnaire was distributed to International Society of Blood Transfusion members in 95 countries. We recorded respondents' demographic information, impacts on the blood supply, CCP collection and use, transfusion demands and operational challenges. Results: Eighty-two responses from 42 countries, including 24 low- and middle-income countries, were analysed. Participants worked in national (26.8%) and regional (26.8%) blood establishments and hospital-based (42.7%) institutions. CCP collection and transfusion were reported by 63% and 36.6% of respondents, respectively. Decreases in blood donations occurred in 70.6% of collecting facilities. Despite safety measures and recruitment strategies, donor fear and refusal of institutions to host blood drives were major contributing factors. Almost half of respondents working at transfusion medicine services were from large hospitals with over 10,000 red cell transfusions per year, and 76.8% of those hospitals experienced blood shortages. Practices varied in accepting donors for blood or CCP donations after a history of COVID-19 infection, CCP transfusion, or vaccination. Operational challenges included loss of staff, increased workloads and delays in reagent supplies. Almost half of the institutions modified their disaster plans during the pandemic. Conclusion: The challenges faced by blood systems during the COVID-19 pandemic highlight the need for guidance, harmonization, and strengthening of the preparedness and the capacity of blood systems against future infectious threats

Clusters of SARS-CoV-2 Lineage B.1.1.7 Infection after Vaccination with Adenovirus-Vectored and Inactivated Vaccines

A SARS-CoV-2 B.1.1.7 variant of concern (VOC) has been associated with increased transmissibility, hospitalization, and mortality. This study aimed to explore the factors associated with B.1.1.7 VOC infection in the context of vaccination. On March 2021, we detected SARS-CoV-2 RNA in nasopharyngeal samples from 14 of 22 individuals vaccinated with a single-dose of ChAdOx1 (outbreak A, n = 26), and 22 of 42 of individuals with two doses of the CoronaVac vaccine (outbreak B, n = 52) for breakthrough infection rates for ChAdOx1 of 63.6% and 52.4% for CoronaVac. The outbreaks were caused by two independent clusters of the B.1.1.7 VOC. The serum of PCR-positive symptomatic SARS-CoV-2-infected individuals had ~1.8–3.4-fold more neutralizing capacity against B.1.1.7 compared to the serum of asymptomatic individuals. These data based on exploratory analysis suggest that the B.1.1.7 variant can infect individuals partially immunized with a single dose of an adenovirus-vectored vaccine or fully immunized with two doses of an inactivated vaccine, although the vaccines were able to reduce the risk of severe disease and death caused by this VOC, even in the elderly