Aspectos imunogenéticos da imunotolerância na gestação e transplantes

O estudo da imunologia reprodutiva data da década de 1950. Seus estudos emergiram das observações pioneiras de Peter Medawar sobre a existência de antígenos de tecido, mais especificamente, em como antígenos expressos em enxertos de pele eram reconhecidos e rejeitados quando transplantados em indivíduos geneticamente distintos. Na gestação, a aceitação do feto é um evento único, e, demonstra como o sistema imunológico materno se adapta e tolera as células fetais semi-alogênicas sem rejeitá-las, mesmo considerando que o feto herda e expressa metade dos antígenos paternos e metade maternos. Diante disso, sugere-se que muitas desordens gestacionais, tais como, a pré-eclâmpsia e abortamentos, sejam causadas por desequilíbrios na tolerância materna. Tal tolerância é alcançada, entre outros mecanismos, através da elevada expressão das moléculas do MHC de classe I não clássico (HLA-E, -F e -G) e ausência de expressão de MHC de classe I clássico (exceto a baixa expressão de HLA-C) e de moléculas de MHC-II, as quais conferem baixa antigenicidade às células fetais. No contexto dos transplantes (histocompatibilidade), a tolerância imunológica é estabelecida de maneira mais complexa, uma vez que deve se considerar dois genomas distintos. A rejeição do transplante é um fenômeno "criado pelo homem", uma vez que o transplante de órgãos não é evidenciado no mundo natural. Embora a rejeição seja altamente dependente da incompatibilidade entre os diferente alelos de HLA entre os pares de doador e receptor, a rejeição aguda ou crônica pode ocorrer mesmo em pares compatíveis, sugerindo a influência de outras moléculas não clássicas do MHC-I e moléculas não-HLA no desfecho dos transplantes. Apesar de serem fenômenos biológicos distintos, as semelhanças entre a gravidez e o transplante não podem ser negligenciadas, especificamente, em relação ao papel imunorregulatório das moléculas do MHC-I não clássico e moléculas não HLA. Na presente tese, apresentaremos o papel das variantes genéticas em genes não clássicos do MHC-I (HLA-E, HLA-G, MIC-A) e receptores cognatos (NKG2C e NKG2D) no contexto da gravidez saudável ou patológica. Para complementar esta tese, avaliamos como os eixos imunológicos NKG2C/HLA-E e NKGD2/MIC-A influenciam na manutenção e sobrevivência do transplante. No total, três artigos publicados em periódicos internacionais e, adicionalmente, três artigos em fase de preparação para publicação compõem está tese. Os resultados desses artigos (publicados) podem ser resumidos como seguem: 1. Michita et al. (2016) A tug-of-war between tolerance and rejection - New evidence for 3'UTR HLAG haplotypes influence in recurrent pregnancy loss (Hum Immunol. 77:892-897). Neste estudo observamos que as variantes genéticas +3010C/G, +3142G/C e +3187A/G localizados na região 3’UTR do HLA-G influenciam na suscetibilidade para o abortamento de repetição numa população brasileira. Além disso, haplótipos observados nessa região foram caracterizados na população estudada, e, a associação do haplótipo UTR-1 com o risco de abortamentos foi observada. 2. Michita et al. (2018) A Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas–Kidney Transplantation Recipients (Int J Mol Sci. 4;19). Nesse estudo avaliamos o impacto da variante MICA Val129Met no desfecho do primeiro ano após o transplante simultâneo de rim e pâncreas. Brevemente, observamos que incompatibilidade no MHC (HLAA, -B, -DR) não influenciaram no desfecho (infecção pelo citomegalovírus, função do órgão e rejeição aguda). Porém, um acentuado efeito no desfecho foi influenciado pela incompatibilidade do alelo MICA 129Val (doador → receptor). Estas observações foram observadas pela primeira vez no contexto de transplantes de órgão sólidos e destacam a importância de outros genes não-HLA no desfecho dos transplantes. 3. Michita et al. (2018) Genetic Variants in Preeclampsia: Lessons From Studies in Latin-American populations (Front Physiol. 4;9:1771). Nesse estudo foi realizado um extensa revisão da literatura no que se refere aos estudos genéticos – variantes genéticas – realizados em populações latinoamericanas investigando a patogênese da pré-eclâmpsia. Este artigo de revisão representa uma importante fonte de dados para estudos futuros devido a sua abrangência e profundidade. Além disso, o artigo destaca os principais grupos engajados na caracterização das bases genéticas da pré-eclâmpsia. Os resultados da presente tese, destacam que apesar das diferenças entre os transplantes e a gestação, diversos mecanismos imunorregulatórios podem se sobrepor nestes dois fenômenos biológicos. Principalmente, no que se refere a investigação das moléculas não clássicas do MHC de classe-I.The field of reproductive immunology dates from the 1950s. It emerged from the pioneering observations of Peter Medawar on tissue antigens, more specifically, in how skin grafts were recognized and rejected when transplanted to a genetically different individual. In pregnancy, the acceptance of the fetus is a unique event, and shows how the maternal immune system should shape to tolerate and do not reject the semi-allogeneic fetal cells, even considering that the fetus inherited half of antigens from each parental - mother and father. Not surprisingly, many pregnancy disorders such as preeclampsia and miscarriages are thought to originate from a breakdown of tolerance. Such tolerance is achieved, amongst other mechanisms, by a high expression of non-classical MHC class I molecules such as HLA-E, -F and –G, and the lack of expression of classical MHC-I (or low expression of HLA-C) and MHC-II molecules. Both such mechanisms allow poor antigenicity to fetal cells. In the context of allotransplantation, immunological tolerance is achieved in a more complex way (histocompatibility), as the attention must be directed to antigens derived from more than one genome. Graft rejection is a “man-made” phenomenon since tissue transplantation is no evidenced in the natural world. Although rejection is highly dependent on HLA mismatches between donor and recipient pairs, even fully matched patients can undergo acute or chronic rejection, suggesting the existence of other non-classical MHC-I and non-HLA molecules driving transplantation outcome. Despite being different biological phenomena, the similarities between pregnancy and transplantation cannot be neglected, is essential to consider the overlapped functions towards a tolerogenic phenotype conferred by non-classical MHC-I and non-MHC molecules. In the current thesis, we will present the role of functional genetic variants in non-classical MHC-I (HLA-E, HLA-G, MIC-A) and cognate receptors (NKG2A and NKG2D) in the context of a healthy or a pathological pregnancy. In order to complement this thesis, we evaluated how the immunological axis NKG2C/HLA-E and NKGD2/MIC-A are involved in the maintenance of organ transplantation. In total, three articles published in international journals and, additionally, three articles in preparation compose the current thesis. The results of these (published) articles can be summarized as follows: 1. Michita et al. (2016) A tug-of-war between tolerance and rejection - New evidence for 3'UTR HLAG haplotypes influence recurrent pregnancy loss (Hum Immunol 77: 892- 897). In this study, we observed that the genetic variants +3010C/G, +3142G/C and +3187A/G located in the 3'UTR region of HLA-G influence the susceptibility to recurrent pregnancy loss in a Brazilian population. In addition, haplotypes observed in this region were characterized in the population, and an association of the UTR-1 haplotype with the risk of miscarriages was observed. 2. Michita et al. (2018) Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas-Kidney Transplantation Recipients (Int J Mol Sci. 4;19). In this study, we evaluated the impact of the MICA Val129Met variant on the first year outcome after simultaneous transplantation of the kidney and pancreas. Briefly, we found that MHC mismatch (HLA-A, -B, -DR) did not influence the outcome (cytomegalovirus infection, organ function, and acute rejection). However, a pronounced effect on the outcome was influenced by the incompatibility of the MICA 129Val allele (donor → recipient). These observations were first observed in the context of solid organ transplants and highlight the importance of other non-HLA genes in the outcome of transplants. 3. Michita et al. (2018) Genetic Variants in Preeclampsia: Lessons From Studies in Latin American Populations (Front Physiol. 4: 9: 1771). In this study, an extensive review of the literature was carried out regarding genetic studies - genetic variants - carried out in Latin American populations investigating the pathogenesis of preeclampsia. This review article represents an essential source of data for future studies because of its breadth and depth. In addition, the article highlights the main groups engaged in characterizing the genetic basis of preeclampsia in Latin America. The results of this thesis point out that despite the differences between transplants and gestation, several immunoregulatory mechanisms can overlap in these two biological phenomena. Mainly for the investigation of non-classical MHC class I molecules

Exploring potential impacts of pregnancy-related maternal immune activation and extracellular vesicles on immune alterations observed in autism spectrum disorder

Autism Spectrum Disorder (ASD) is a set of neurodevelopmental disorders usually observed in early life, with impacts on behavioral and social skills. Incidence of ASD has been dramatically increasing worldwide, possibly due to increase in awareness/diagnosis as well as to genetic and environmental triggers. Currently, it is estimated that ~1% of the world population presents ASD symptoms. In addition to its genetic background, environmental and immune-related factors also influence the ASD etiology. In this context, maternal immune activation (MIA) has recently been suggested as a component potentially involved in ASD development. In addition, extracellular vesicles (EVs) are abundant at the maternal-fetal interface and are actively involved in the immunoregulation required for a healthy pregnancy. Considering that alterations in concentration and content of EVs have also been associated with ASD, this article raises a debate about the potential roles of EVs in the processes surrounding MIA. This represents the major differential of the present review compared to other ASD studies. To support the suggested correlations and hypotheses, findings regarding the roles of EVs during pregnancy and potential influences on ASD are discussed, along with a review and update concerning the participation of infections, cytokine unbalances, overweight and obesity, maternal anti-fetal brain antibodies, maternal fever, gestational diabetes, preeclampsia, labor type and microbiota unbalances in MIA and ASD

Helicobacter pylori eradication : influence of interleukin-1beta –31 C/T polymorphism

Aim: To analyze the influence of the –31 C/T polymorphism of the interleukin-1 gene on Helicobacter pylori eradication therapy success in patients with functional dyspepsia. Methods: Functional dyspepsia was diagnosed according to the Rome III criteria. All patients underwent upper gastrointestinal endoscopy, and gastric biopsies were obtained at screening and 12 months after randomization (last follow-up visit). Urease test and histological examination were performed to define the H. pylori status. Patients received twice-daily amoxicillin, clarithromycin and omeprazole for 10 days. Genotyping of the interleukin-1beta –31 C/T polymorphism (rs1143627)was performed using polymerase chain reaction-restriction fragment length polymorphism. Results: One hundred forty-nine patients received treatment with triple therapy for H. pylori eradication. Only one patient was lost to follow-up, and adherence to study medication was 94.6%. A total of 148 patients (mean age 46.08±12.24 years; 81.8% women) were evaluated for the influence of the interleukin-1beta –31 C/T polymorphism on the outcome of H. pylori eradication therapy. After treatment, bacteria were eradicated in 87% of patients (129/148). Genotype frequencies of the polymorphism were as follows: CC, 38/148 (25.7%); CT, 71/148 (47.9%); and TT, 39/148 (26.4%). Successful eradication rate was 78.9%, 94.4% and 82.1% for the CC, CT and TT genotypes, respectively. The CT genotype was significantly associated with successful H. pylori eradication (p = 0.039). Conclusion: This study suggests that the CT genotype of the interleukin-1beta –31 C/T polymorphism plays a role in the successful eradication of H. pylori among patients with functional dyspepsia

Apoptose na gestação : análise de variantes polimórficas dos genes FAS, FAS-L, BAX, BCL-2 e HLA-G na etiologia do abortamento de repetição

O Abortamento de repetição (AR) é uma condição patológica definida pela ocorrência de duas ou mais perdas gestacionais consecutivas. Estima-se que esta desordem acometa 5% dos casais em idade reprodutiva. Esta condição clínica é classificada em dois subtipos clínicos: AR primário e AR secundário. Apesar dos esforços em investigar as causas desta desordem obstétrica, estima-se que em 50% dos casos a etiologia do AR permanece desconhecida. A reprodução humana implica em um paradoxo imunológico fundamental: no qual o feto representa uma entidade estranha ao sistema imune materno, sendo constituído por metade de material genético paterno, no entanto, de forma fascinante este não é rejeitado. Sendo assim, a aceitação materna do feto é um evento único e demonstra como o sistema imune materno remodela-se e tolera a presença de células invasivas semi-alogênicas no útero. Casos de AR, pré-eclâmpsia (PE), entre outras desordens gestacionais, levam a uma questão retórica; ‘Porque a mãe rejeitou o feto?’. No entanto, considerando as gestações saudáveis e as complexas interações que ocorrem na interface materno-placentária, talvez, a intrigante e fascinante questão correta que ainda permanece é ‘Porque a mãe não rejeitou o feto?’. Existem muitos mecanismos que protegem o feto de um possível ‘ataque’ do sistema imune materno das quais podemos citar: (i) a baixa expressão de moléculas clássicas de classe I do complexo principal de histocompatibilidade (MHC) na interface materno-placentária, (ii) a expressão diferencial de moléculas não clássicas de classe I do MHC nas células do trofoblasto, como HLA (antígeno leucocitário humano)-E, HLA-F e HLA-G, (iii) a apoptose de células do sistema imunológico ativadas, entre outras. A morte celular programada ou apoptose é essencial ao desenvolvimento e funcionalidade dos organismos multicelulares. Os mecanismos apoptóticos mais reconhecidos pela efetividade em desencadear a morte celular são: o sistema FAS-FAS-L (via extrínseca) e BAX-BCL-2 (via intrínseca). A molécula do HLA-G, descrita por sua versatilidade imunológica, é essencial na gestação e neste cenário é descrito que, além de contribuir essencialmente na imunotolerância materna, também atua nos processos de angiogênese, apoptose e consequente remodelação tecidual da decídua materna. Considerando que os eventos apoptóticos estão presentes nas diversas etapas da gestação, desde a concepção até o nascimento, e que constituem diferentes interações de proteínas apoptóticas e anti-apoptóticas específicas para manter a adequada homeostasia celular e tecidual, o presente estudo visa: (i) a compreensão dos principais genes envolvidos neste processo e (ii) o entendimento da contribuição destes na predisposição à recorrência de abortamentos, em um estudo de carácter caso-controle. No total, 138 mulheres diagnosticadas com AR e 156 mulheres saudáveis com mais de dois filhos, sem histórico de complicações gestacionais foram incluídas neste estudo. Os genes e variantes polimórficas analisados no estudo a menos diferentemente especificado correspondem a variações presente na região promotora, são: FAS (rs1800682, rs763110), FASL-L (rs5030772, rs763110), BAX (rs4645878), BCL-2 (rs2279115) e HLA-G [região 3̍ UTR (região não traduzida) (rs66554220, rs1707, rs1710, rs17179101, rs17179108, rs1063320, rs9380142 e rs1610696)]. Como conclusão, no presente estudo nós não observamos a associação destas variantes ao AR na população estudada. No entanto, algumas ressalvas a respeito deste estudo devem ser mencionadas, são estas: (i) a heterogeneidade genética da população estudada, (ii) o tamanho amostral e (iii) o critério clínico para o diagnóstico do abortamento de repetição como 2 (duas) ou mais perdas gestacionais consecutivas. Portanto, o papel destas variantes genéticas na predisposição ao AR não deve ser descartada e necessitam ser confirmadas em estudos posteriores. Como perspectivas, no intuito de gerar evidências mais satisfatórias sobre o ofício destas variantes genéticas na fisiopatologia do AR, estudos com adequado tamanho amostral capaz de detectar tais efeitos funcionais destas variantes, bem como em diferentes populações são fortemente recomendados.Recurrent miscarriage (RM) or recurrent pregnancy loss (RPL) is a pathological condition defined by occurrence of two or more consecutive pregnancy losses. It is estimated that about 5% of couples in reproductive age are affected by RM. This clinical condition is classified in two subtypes: primary and secondary RM. In despite of the efforts to investigate the causes of RM, approximately 50% of cases the etiology of RM remains unknown. Human reproduction involves a fundamental immunological paradox: in which the fetus is seem as foreign entity to the maternal immune system, consisting of half maternal and half paternal genetic material origin; however, in a fascinating way the fetus is not rejected. Thus, maternal-fetal acceptance is a unique example and demonstrates how the maternal immune system shapes and tolerates up the presence of invasive semi-allogeneic cells in utero. Obstetric disorders such as RM, preeclampsia (PE) and other pregnancy complications, lead to a rhetorical question: 'Why the mother rejected the fetus?'. However, considering all healthy pregnancies and complex interactions that occur in the maternalplacental interface, perhaps the intriguing question that remains is 'why the mother did not reject the fetus?’ there are many mechanisms that protect the fetus of a possible 'attack' from maternal immune system. Among them we can cite: (i) the low expression of classical class I molecules of the major histocompatibility complex (MHC) in maternalplacental interface, (ii) the differential expression of non-classical class I molecules MHC on the trophoblast cells such as HLA (human leukocyte antigen)-E, HLA-F and HLA-G, (iii) tryptophan catabolism by the enzyme indoleamine (2,3)-dioxigenase, (iv) regulation of the complement system in maternal-placental interface and (v) apoptosis of maternal activated immune system cells. Programmed cell death or apoptosis is essential to development and function of multicellular organisms. Apoptotic mechanisms recognized for the effectiveness in triggering cell death are: FAS-FAS-L system (extrinsic pathway) and BAX-BCL-2 (intrinsic pathway). HLA-G molecule is described by its immunological versatility being essential to establishment of a healthy gestation. The HLA-G contributes primarily to maternal immunotolerance and is related to many process, such as angiogenesis, apoptosis and consequently to tissue remodeling of maternal decidua. Since the apoptotic events are present at different stages of pregnancy, from conception to birth, and considering different interactions of apoptotic and specific anti-apoptotic proteins to maintain a suitable cellular and tissue homeostasis, this study aims to: (i) the understanding of key genes involved in the apoptotic process and (ii) the understanding of these genes in the predisposition to recurrent miscarriages, in a case-control study. A total of 138 women diagnosed with RM and 156 healthy women with more than two children with no history of pregnancy complications were included in this study. All polymorphic variants of candidate genes analyzed in the study unless otherwise specified correspond to variations in the promoter region. The variants assessed in study are: FAS (rs1800682, rs763110), FASL-G (rs5030772, rs763110), BAX (rs4645878), BCL-2 (rs2279115) and HLA-G [region 3̍ UTR (untranslated region) (rs66554220, rs1707, rs1710, rs17179101, rs17179108, rs1063320, rs9380142 and rs1610696)]. In conclusion, we did not observe the association of these variants to the RM risk in the present population. However, some caveats about this study should be mentioned, such as (i) the genetic heterogeneity of the study population, (ii) the sample size and (iii) the clinical criteria for the diagnosis of recurrent miscarriage as two (2) or more consecutive pregnancy losses. Therefore, a definitive conclusion on the role of these genetic variants in the predisposition to RM should not be dismissed and should be further confirmed in future studies. As prospects to generate more satisfactory evidence about the role of these genetic variants in the pathophysiology of RM, further studies with adequate sample size capable of detecting such functional effects of these variants as well as in different populations are strongly recommended

A Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas–Kidney Transplantation Recipients

The polymorphic major histocompatibility complex class I chain-related molecule A (MICA) and its soluble form (sMICA) interact with activating receptor natural-killer group 2 member D (NKG2D) on natural-killer (NK) and T cells, thereby modifying immune responses to transplantation and infectious agents (e.g., cytomegalovirus). Two single-nucleotide polymorphisms (SNPs), rs2596538GA in the MICA promoter and rs1051792AG in the coding region (MICA-129Val/Met), influence MICA expression or binding to NKG2D, with MICA-129Met molecules showing higher receptor affinity. To investigate the impact of these SNPs on the occurrence of cytomegalovirus infection or acute rejection (AR) in individuals who underwent simultaneous pancreas–kidney transplantation (SPKT), 50 recipient-donor pairs were genotyped, and sMICA levels were measured during the first year post-transplantation. Recipients with a Val-mismatch (recipient Met/Met and donor Val/Met or Val/Val) showed shorter cytomegalovirus infection-free and shorter kidney AR-free survival. Additionally, Val mismatch was an independent predictor of cytomegalovirus infection and kidney AR in the first year post-transplantation. Interestingly, sMICA levels were lower in rs2596538AA and MICA129Met/Met-homozygous recipients. These results provide further evidence that genetic variants of MICA influence sMICA levels, and that Val mismatch at position 129 increases cytomegalovirus infection and kidney AR risk during the first year post-SPKT

Aspectos imunogenéticos da imunotolerância na gestação e transplantes

O estudo da imunologia reprodutiva data da década de 1950. Seus estudos emergiram das observações pioneiras de Peter Medawar sobre a existência de antígenos de tecido, mais especificamente, em como antígenos expressos em enxertos de pele eram reconhecidos e rejeitados quando transplantados em indivíduos geneticamente distintos. Na gestação, a aceitação do feto é um evento único, e, demonstra como o sistema imunológico materno se adapta e tolera as células fetais semi-alogênicas sem rejeitá-las, mesmo considerando que o feto herda e expressa metade dos antígenos paternos e metade maternos. Diante disso, sugere-se que muitas desordens gestacionais, tais como, a pré-eclâmpsia e abortamentos, sejam causadas por desequilíbrios na tolerância materna. Tal tolerância é alcançada, entre outros mecanismos, através da elevada expressão das moléculas do MHC de classe I não clássico (HLA-E, -F e -G) e ausência de expressão de MHC de classe I clássico (exceto a baixa expressão de HLA-C) e de moléculas de MHC-II, as quais conferem baixa antigenicidade às células fetais. No contexto dos transplantes (histocompatibilidade), a tolerância imunológica é estabelecida de maneira mais complexa, uma vez que deve se considerar dois genomas distintos. A rejeição do transplante é um fenômeno "criado pelo homem", uma vez que o transplante de órgãos não é evidenciado no mundo natural. Embora a rejeição seja altamente dependente da incompatibilidade entre os diferente alelos de HLA entre os pares de doador e receptor, a rejeição aguda ou crônica pode ocorrer mesmo em pares compatíveis, sugerindo a influência de outras moléculas não clássicas do MHC-I e moléculas não-HLA no desfecho dos transplantes. Apesar de serem fenômenos biológicos distintos, as semelhanças entre a gravidez e o transplante não podem ser negligenciadas, especificamente, em relação ao papel imunorregulatório das moléculas do MHC-I não clássico e moléculas não HLA. Na presente tese, apresentaremos o papel das variantes genéticas em genes não clássicos do MHC-I (HLA-E, HLA-G, MIC-A) e receptores cognatos (NKG2C e NKG2D) no contexto da gravidez saudável ou patológica. Para complementar esta tese, avaliamos como os eixos imunológicos NKG2C/HLA-E e NKGD2/MIC-A influenciam na manutenção e sobrevivência do transplante. No total, três artigos publicados em periódicos internacionais e, adicionalmente, três artigos em fase de preparação para publicação compõem está tese. Os resultados desses artigos (publicados) podem ser resumidos como seguem: 1. Michita et al. (2016) A tug-of-war between tolerance and rejection - New evidence for 3'UTR HLAG haplotypes influence in recurrent pregnancy loss (Hum Immunol. 77:892-897). Neste estudo observamos que as variantes genéticas +3010C/G, +3142G/C e +3187A/G localizados na região 3’UTR do HLA-G influenciam na suscetibilidade para o abortamento de repetição numa população brasileira. Além disso, haplótipos observados nessa região foram caracterizados na população estudada, e, a associação do haplótipo UTR-1 com o risco de abortamentos foi observada. 2. Michita et al. (2018) A Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas–Kidney Transplantation Recipients (Int J Mol Sci. 4;19). Nesse estudo avaliamos o impacto da variante MICA Val129Met no desfecho do primeiro ano após o transplante simultâneo de rim e pâncreas. Brevemente, observamos que incompatibilidade no MHC (HLAA, -B, -DR) não influenciaram no desfecho (infecção pelo citomegalovírus, função do órgão e rejeição aguda). Porém, um acentuado efeito no desfecho foi influenciado pela incompatibilidade do alelo MICA 129Val (doador → receptor). Estas observações foram observadas pela primeira vez no contexto de transplantes de órgão sólidos e destacam a importância de outros genes não-HLA no desfecho dos transplantes. 3. Michita et al. (2018) Genetic Variants in Preeclampsia: Lessons From Studies in Latin-American populations (Front Physiol. 4;9:1771). Nesse estudo foi realizado um extensa revisão da literatura no que se refere aos estudos genéticos – variantes genéticas – realizados em populações latinoamericanas investigando a patogênese da pré-eclâmpsia. Este artigo de revisão representa uma importante fonte de dados para estudos futuros devido a sua abrangência e profundidade. Além disso, o artigo destaca os principais grupos engajados na caracterização das bases genéticas da pré-eclâmpsia. Os resultados da presente tese, destacam que apesar das diferenças entre os transplantes e a gestação, diversos mecanismos imunorregulatórios podem se sobrepor nestes dois fenômenos biológicos. Principalmente, no que se refere a investigação das moléculas não clássicas do MHC de classe-I.The field of reproductive immunology dates from the 1950s. It emerged from the pioneering observations of Peter Medawar on tissue antigens, more specifically, in how skin grafts were recognized and rejected when transplanted to a genetically different individual. In pregnancy, the acceptance of the fetus is a unique event, and shows how the maternal immune system should shape to tolerate and do not reject the semi-allogeneic fetal cells, even considering that the fetus inherited half of antigens from each parental - mother and father. Not surprisingly, many pregnancy disorders such as preeclampsia and miscarriages are thought to originate from a breakdown of tolerance. Such tolerance is achieved, amongst other mechanisms, by a high expression of non-classical MHC class I molecules such as HLA-E, -F and –G, and the lack of expression of classical MHC-I (or low expression of HLA-C) and MHC-II molecules. Both such mechanisms allow poor antigenicity to fetal cells. In the context of allotransplantation, immunological tolerance is achieved in a more complex way (histocompatibility), as the attention must be directed to antigens derived from more than one genome. Graft rejection is a “man-made” phenomenon since tissue transplantation is no evidenced in the natural world. Although rejection is highly dependent on HLA mismatches between donor and recipient pairs, even fully matched patients can undergo acute or chronic rejection, suggesting the existence of other non-classical MHC-I and non-HLA molecules driving transplantation outcome. Despite being different biological phenomena, the similarities between pregnancy and transplantation cannot be neglected, is essential to consider the overlapped functions towards a tolerogenic phenotype conferred by non-classical MHC-I and non-MHC molecules. In the current thesis, we will present the role of functional genetic variants in non-classical MHC-I (HLA-E, HLA-G, MIC-A) and cognate receptors (NKG2A and NKG2D) in the context of a healthy or a pathological pregnancy. In order to complement this thesis, we evaluated how the immunological axis NKG2C/HLA-E and NKGD2/MIC-A are involved in the maintenance of organ transplantation. In total, three articles published in international journals and, additionally, three articles in preparation compose the current thesis. The results of these (published) articles can be summarized as follows: 1. Michita et al. (2016) A tug-of-war between tolerance and rejection - New evidence for 3'UTR HLAG haplotypes influence recurrent pregnancy loss (Hum Immunol 77: 892- 897). In this study, we observed that the genetic variants +3010C/G, +3142G/C and +3187A/G located in the 3'UTR region of HLA-G influence the susceptibility to recurrent pregnancy loss in a Brazilian population. In addition, haplotypes observed in this region were characterized in the population, and an association of the UTR-1 haplotype with the risk of miscarriages was observed. 2. Michita et al. (2018) Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas-Kidney Transplantation Recipients (Int J Mol Sci. 4;19). In this study, we evaluated the impact of the MICA Val129Met variant on the first year outcome after simultaneous transplantation of the kidney and pancreas. Briefly, we found that MHC mismatch (HLA-A, -B, -DR) did not influence the outcome (cytomegalovirus infection, organ function, and acute rejection). However, a pronounced effect on the outcome was influenced by the incompatibility of the MICA 129Val allele (donor → recipient). These observations were first observed in the context of solid organ transplants and highlight the importance of other non-HLA genes in the outcome of transplants. 3. Michita et al. (2018) Genetic Variants in Preeclampsia: Lessons From Studies in Latin American Populations (Front Physiol. 4: 9: 1771). In this study, an extensive review of the literature was carried out regarding genetic studies - genetic variants - carried out in Latin American populations investigating the pathogenesis of preeclampsia. This review article represents an essential source of data for future studies because of its breadth and depth. In addition, the article highlights the main groups engaged in characterizing the genetic basis of preeclampsia in Latin America. The results of this thesis point out that despite the differences between transplants and gestation, several immunoregulatory mechanisms can overlap in these two biological phenomena. Mainly for the investigation of non-classical MHC class I molecules

Recipient HLA-G +3142 CC Genotype and Concentrations of Soluble HLA-G Impact on Occurrence of CMV Infection after Living-Donor Kidney Transplantation

The expression modulation of the immunosuppressive non-classical Human leukocyte antigen-G (HLA-G) molecule and its soluble isoforms is an immune evasion strategy being deployed by cytomegalovirus (CMV). The +3142 C>G single nucleotide polymorphism (SNP) located within the 3′ untranslated region (3′UTR) is of crucial importance for the regulation of HLA-G expression. Therefore, we analyzed the influence of the +3142 C>G HLA-G SNP on the occurrence of CMV infection in a cohort of 178 living-donor kidney recipients and their 178 corresponding donors. In addition, soluble HLA-G (sHLA-G) levels were quantified before and after transplantation. The presence of the HLA-G +3142 CC genotype in recipients, but not donors of our cohort as along with elevated sHLA-G levels (≥ 6.1 ng/mL) were associated with higher susceptibility to CMV infection after transplantation. Our results provided evidence that i) HLA-G is implicated in the establishment of CMV after living-donor kidney transplantation and ii) recipient HLA-G +3142 CC genotype and sHLA-G concentration levels could represent important predictive risk markers for CMV infection

A Valine Mismatch at Position 129 of MICA Is an Independent Predictor of Cytomegalovirus Infection and Acute Kidney Rejection in Simultaneous Pancreas–Kidney Transplantation Recipients

The polymorphic major histocompatibility complex class I chain-related molecule A (MICA) and its soluble form (sMICA) interact with activating receptor natural-killer group 2 member D (NKG2D) on natural-killer (NK) and T cells, thereby modifying immune responses to transplantation and infectious agents (e.g., cytomegalovirus). Two single-nucleotide polymorphisms (SNPs), rs2596538GA in the MICA promoter and rs1051792AG in the coding region (MICA-129Val/Met), influence MICA expression or binding to NKG2D, with MICA-129Met molecules showing higher receptor affinity. To investigate the impact of these SNPs on the occurrence of cytomegalovirus infection or acute rejection (AR) in individuals who underwent simultaneous pancreas–kidney transplantation (SPKT), 50 recipient-donor pairs were genotyped, and sMICA levels were measured during the first year post-transplantation. Recipients with a Val-mismatch (recipient Met/Met and donor Val/Met or Val/Val) showed shorter cytomegalovirus infection-free and shorter kidney AR-free survival. Additionally, Val mismatch was an independent predictor of cytomegalovirus infection and kidney AR in the first year post-transplantation. Interestingly, sMICA levels were lower in rs2596538AA and MICA129Met/Met-homozygous recipients. These results provide further evidence that genetic variants of MICA influence sMICA levels, and that Val mismatch at position 129 increases cytomegalovirus infection and kidney AR risk during the first year post-SPKT