O requisito fundamental na seleção do par doador-receptor em Transplantes de Medula Óssea (TMO) é regido pelo sistema do Complexo Principal de Histocompatibilidade, ou seja, pelos mecanismos imunológicos mediados pelas moléculas dos Antígenos Leucocitários Humanos (HLA). No entanto as incompatibilidades HLA, podem influenciar de forma negativa ou positiva os resultados dos transplantes, através da Doença do Enxerto versus Hospedeiro e o efeito do enxerto versus Leucemia (EvL) respectivamente. Ainda é desconhecido o impacto do locus HLA-DPB1* neste contexto. Assim o presente projeto tem como objetivo a avaliação do impacto do HLA-DPB1* em transplantes de pacientes consanguíneos e a ocorrência de DECH. Para a tal finalidade, tipificamos o locus em questão utilizando a metodologia PCR-SSO, onde após a reação de amplificação da cadeia pela polimerase, realizamos a hibridização com uma sequência específica de oligonucleotídeos para tipificação do Loco HLA-DPB1*. Foram analisadas 826 amostras, sendo 413 pares de receptores e seus respectivos doadores familiares, submetidos a Transplantes de Células Tronco Hematopoiéticas, realizados na Unidade de Transplante de Medula Óssea de Curitiba da Universidade Federal do Paraná e da Faculdade de Medicina de Ribeirão Preto- USP. Observou-se que a presença de incompatibilidades HLA-DPB1* aumentam a chance dos receptores desenvolverem a doença do enxerto versus hospedeiro aguda, em graus mais graves. Assim, concluímos que a avaliação deste loco pode prevenir esta doença, e caso não haja outro doador, alerta o clínico quanto à utilização de medidas profiláticas. / The key requirement in the selection of the receptor-donor pair for bone marrow transplant is is defined by the Major Histocompatibility Complex, or by immunologic mechanisms mediated by molecules of the Human Leukocyte Antigens (HLA). However the post transplant complications due to HLA mismatches, as Graft versus Host Disease (GVHD) and graft failure are fundamental to the success of these transplants. Still unknown is the impact of loci HLA DPB1*, so this project aims to assess the impact of HLA - DPB1* in transplant patients consanguineous and assessing the impact of incompatibilities in HLA - DPB1 * GVHD. For this purpose, analyzed the loco in question using the PCR-SSO method, where after the amplification reaction polymerase chain, we performed hybridization with a sequence -specific primers for typing of HLA - DPB1* Loco. We analyzed 826 samples, 413 pairs of recipients and their respective donors, patients undergoing Hematopoietic Stem Cell Transplants performed in the Unit for Bone Marrow Transplantation in Curitiba, Federal University of Paraná and the Faculty of Medicine of Ribeirão Preto - USP. It was observed that the presence of mismatches HLA- DPB1* increase the chance of recipients develop chronic graft versus host disease, in more severe degrees. Thus, we conclude that the evaluation of this loci can prevent this disease and if no other donors alert the clinician to the use of prophylactic measures.

Submitted by Erika Demachki (erikademachki@gmail.com) on 2014-11-11T20:37:41Z No. of bitstreams: 2 Dissertação - Leandro Prado Felício - 2013.pdf: 4839350 bytes, checksum: 07898140311917429ab55aa63ad5324e (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Erika Demachki (erikademachki@gmail.com) on 2014-11-11T20:38:05Z (GMT) No. of bitstreams: 2 Dissertação - Leandro Prado Felício - 2013.pdf: 4839350 bytes, checksum: 07898140311917429ab55aa63ad5324e (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-11-11T20:38:05Z (GMT). No. of bitstreams: 2 Dissertação - Leandro Prado Felício - 2013.pdf: 4839350 bytes, checksum: 07898140311917429ab55aa63ad5324e (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2013-01-31 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / The HLA-E locus is a Human Major Histocompatibility Complex (MHC) gene associated with immune-modulation and suppression of the immune response by the interaction with specific NK and T cell receptors. The HLA-E gene is considered the most conserved locus in the human HLA; however, this low variability might be a consequence of the scarce number of studies focusing this subject. In this mastering thesis we assessed the HLA-E coding and 3’ untranslated region variability in a group of individuals from Brazil and the results were evaluated together with data from the 1000Genomes Consortium. Altogether, only 28 variation sites were found in approximately 2724 bp evaluated. These variation sites were arranged into 33 haplotypes, most of them (98.2%) encoding one of the two HLA-E molecules found worldwide, i.e., the molecules associated with the allele groups E*01:01 and E*01:03. Interestingly, 85% of all haplotypes were represented by only three different sequences, each of them associated with one of the main known HLA-E coding alleles, E*01:01:01, E*01:03:01 and E*01:03:02, all of them found worldwide. This phenomenon, together with the comparisons with other primate sequences, reveals that these two main allele groups (and molecules) arose early before human speciation, and indicates that E*01:03:01 might be the oldest allele. In addition, the low nucleotide diversity found for the HLA-E coding and 3’UTR in worldwide populations suggests that the HLA-E gene is in fact a conserved gene, which might be a consequence of its key role in the modulation of the immune system. / O loco HLA-E é um gene do Complexo Principal de Histocompatibilidade Humano (MHC), cujo produto está relacionado com a modulação e supressão da resposta imunitária por meio da interação com receptores específicos das células NK e linfócitos T. O gene HLA-E é considerado o loco menos polimórfico dos genes do complexo HLA, no entanto, esta baixa variabilidade pode ser uma consequência do pequeno número de estudos realizados sobre esse tema. No presente trabalho, a variabilidade das regiões codificadoras e 3’ não traduzida do gene HLA-E foi analisada em amostras brasileiras e os resultados foram comparados com dados obtidos pelo projeto 1000Genomes. Considerando todas as populações avaliadas, apenas 28 pontos de variação foram encontrados em uma região de aproximadamente 2724-pb. Estes pontos de variação estão arranjados em 33 haplótipos diferentes, a maioria deles (98%) codificando uma das duas moléculas HLA-E frequentemente encontradas, E*01:01 e E*01:03. Ainda, 85% dos haplótipos encontrados foram representados por apenas três sequências diferentes, cada uma deles associada a um dos principais alelos da região codificadora do gene HLA-E, E*01:01:01, E*01:03:01 e E*01:03:02. Todas essas sequências foram encontradas em todas as populações avaliadas. Este fenômeno, em conjunto com as comparações envolvendo sequências de primatas, sugere que estes dois grupos de alelos principais (e moléculas) surgiram antes da especiação e dispersão humana, além de indicar que o alelo E*01:03:01 pode ser o mais antigo dentre os demais. Ainda, a baixa diversidade nucleotídica encontrada para a região codificadora e 3' NT do gene HLA-E em populações de todo o mundo sugere que este gene é, de fato, bastante conservado, provavelmente devido ao seu papel chave na modulação das respostas imunes.

Submitted by Franciele Moreira (francielemoreyra@gmail.com) on 2017-09-13T17:35:41Z No. of bitstreams: 2 Dissertação - Mariana Paiva Lopes - 2014 .pdf: 2192084 bytes, checksum: e551c174d821b3b398247680a94c40cd (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-09-19T13:59:44Z (GMT) No. of bitstreams: 2 Dissertação - Mariana Paiva Lopes - 2014 .pdf: 2192084 bytes, checksum: e551c174d821b3b398247680a94c40cd (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-09-19T13:59:44Z (GMT). No. of bitstreams: 2 Dissertação - Mariana Paiva Lopes - 2014 .pdf: 2192084 bytes, checksum: e551c174d821b3b398247680a94c40cd (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2014-03-12 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES / The gene for tumor necrosis factor (TNF) is located in the human MHC central region known as class III (Major Histocompatibility Complex). This gene encodes an important pro- inflammatory cytokine produced primarily by macrophages, and it has been associated with several diseases, such as autoimmune and degenerative ones. Studies indicate that TNF polymorphisms may influence their level of expression and activity and therefore could influence susceptibility to tumors. Whereas the bladder urothelium can constantly be the target of inflammatory processes and as the main forms of treatment of bladder tumors are immunotherapy, the genetic profile of an individual can influence susceptibility to this type of injury. In this study, we analyzed the structure and variability of the regulatory region of the TNF gene in Brazilian samples and the results were compared with data obtained by 1000Genomes project. A study of association between polymorphisms of the promoter region of TNF and bladder carcinoma patients in the state of São Paulo in Ribeirão Preto, in our analysis was conducted there was no relationship of the data found with bladder carcinoma. In all evaluated populations we found 15 variation points, found in 11 Brazilian and 15 variation points found in the 1000Genomes data, these variation points are arranged in 20 different haplotypes. These haplotypes with comparisons involving primate sequences indicated that one allele arose before the main speciation and human dispersion. Two strains were defined by haplotype relationships and are probably very old in human evolutionary history, since all populations evaluated showed haplotypes belonging to each of these strains. The frequency of haplotype H01 is the highest among all populations evaluated. However, the haplotype H12, although at reduced frequency compared to H01 is probably the oldest haplotype in part by the second line being shared with other primates. / O gene do Fator de Necrose Tumoral (TNF) humano está localizado no MHC (Complexo Principal de Histocompatibilidade) central, região conhecida como classe III. Este gene codifica uma citocina pró-inflamatória importante, produzida principalmente por macrófagos, que tem sido relacionada com diversas doenças, como as autoimunes e as degenerativas. Estudos indicam que polimorfismos do TNF podem influenciar seu nível de expressão e atividade e, portanto, poderiam influenciar a susceptibilidade a tumores. Considerando que o urotélio vesical pode ser constantemente alvo de processos inflamatórios e que as principais formas de tratamento de tumores vesicais são imunoterápicos, o perfil genético de um indivíduo pode influenciar a susceptibilidade a esse tipo de lesão. Neste estudo analisamos a estrutura e a variabilidade da região regulatória do gene TNF em amostras brasileiras e os resultados foram comparados com dados obtidos pelo projeto 1000Genomes. Foi realizado um estudo de associação entre polimorfismos da região promotora do TNF e o carcinoma vesical em pacientes do estado de São Paulo da cidade de Ribeirão Preto, nas nossas análises não houve relação dos dados encontrados com o carcinoma vesical. Considerando todas as populações avaliadas foram encontrados 15 pontos de variação, 11 encontrados nas amostras brasileiras e 15 encontrados nos dados do projeto 1000Genomes, esses pontos de variação estão arranjados em 20 haplótipos diferentes. Esses haplótipos em conjunto com as comparações envolvendo sequências de primatas indicam que um alelo principal surgiu antes da especiação e dispersão humana. Duas linhagens foram definidas pelas relações haplotípicas e são, provavelmente, muito antigas na história evolutiva humana, já que todas as populações avaliadas apresentaram haplótipos pertencentes a cada uma destas linhagens. A frequência do haplótipo H01 é a mais alta entre todas as populações avaliadas. No entanto, o haplótipo H12, embora com frequência reduzida quando comparado com o H01, provavelmente é o haplótipo mais antigo em parte por esta segunda linhagem ser compartilhada com outros primatas.

Assessing the genetics of wild animal populations aims to understand selective pressures, and factors whether it be inbreeding or adaptation, that affect the genome. Although numerous techniques are available for assessing population structure, a major obstacle in studying wild populations is obtaining samples from the animals without having to capture them, which can lead to undue distress and injury. Therefore, biologists often use non-invasive sampling methods (i.e., collection of feces, hair) to extract host DNA. In this study, new DNA extraction protocols were developed that improved the quality and quantity of DNA obtained from fecal matter. Fecal samples aged up to Day 6 as well as field samples with unknown days since defecation were successful in individualization of the contributors using microsatellites and were further used to demonstrate kinship. Neutral markers such as short tandem repeat, and mitochondrial D-loop sequences are used for assessing relatedness and evolutionary relationships and can mutate without detrimental effects on the organism. Loci, such as the major histocompatibility complex (MHC), adapt more rapidly under selective pressure such as parasite load, or resistance to diseases and support natural selection processes. Analysis of the neutral microsatellites in Big Summit feral horse population demonstrated a population lacking diversity and trending towards being an inbred population. However, examination of the MHC genes showed maintenance of greater variation that may be the result of selection pressures. The MHC similarity and lower genetic demarcation between geographically separated horse populations further indicated effect of selection pressures in preserving diversity at the MHC genes. Although such molecular markers are used in profiling populations, the current study was also successful in demonstrating the use of individual odor profiles as an additional profiling tool. Volatile organic compounds (VOC) obtained from hair of domestic horses were able to individualize horses as well as differentiate between horse breeds and display kinship. The relation of genetics to odor phenotype is of interest as the inherent polymorphic nature of MHC genes has the potential to generate unique combinations of genotypes that presumably produce distinct odor phenotypes. Subsequently, this study was able to show a significant correlation between MHC genotypes and VOC odor profiles in horses. Understanding the relationship between MHC and odor using domestic horses with known relatedness provides evidence that these same correlations may be applicable to wild equids and dictates their harem hierarchal social structure.

Orientador: Maria Helena Stangler Kraemer / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-14T09:37:26Z (GMT). No. of bitstreams: 1 Magalhaes_RenataFerreira_D.pdf: 45600840 bytes, checksum: c3be46b9fbfab6e5433e2e91db336bb3 (MD5) Previous issue date: 2009 / Resumo: Psoríase é uma dermatose inflamatória crônica, determinada por desregulação do sistema imune e associada a várias comorbidades. O marcador genético mais associado à psoríase em todas as populações é o HLA-CW06. Polimorfismos na região promotora do TNF-a, especialmente a troca de uma guanina por uma adenina nas posições -238 e -308 estão relacionados à alta produção de TNF-a e risco aumentado para psoríase nas populações caucasóides e não em asiáticos. Com o objetivo de determinar se polimorfismos destes genes podem ser fatores de risco para susceptibilidade ou gravidade da doença em pacientes brasileiros, foi realizado um estudo caso-controle com 69 pacientes com psoríase de início até 40 anos, com acompanhamento por dez anos para caracterização de sua evolução clínica em doença leve (grupo I) e grave (grupo II), e 70 indivíduos sadios. Foi feita a identificação dos alelos HLA classe I e II e SNPs da região promotora do TNF-a -238 e -308. Coletaram-se 10 mL de sangue periférico dos indivíduos e se realizou a extração do DNA através do método salting out. O DNA foi amplificado pela reação em cadeia da polimerase (PCR), com primers sequência-específica. As freqüências alélicas e gênicas foram estimadas por contagem direta e a comparação entre as frequências dos grupos foi efetuada por Teste de Fisher (programa GraphPad InStat 3.05). Dois métodos computacionais foram usados para determinar os haplótipos dos indivíduo: (1) o algoritmo ELB implementado pelo software Arlequin 3.1 e (2) um método de base coalescente implementado pelo software PHASE v2, e as freqüências de cada haplótipo foram comparadas por Teste de Fisher. No grupo II, observou-se maior associação com fatores desencadeantes como estresse, início na adolescência e predominância do sexo masculino. Pode-se sugerir que os alelos HLA-B*37, -Cw*06, -Cw*12 e -DRB1*07 foram associados ao curso mais grave da doença, enquanto - B*57 à doença mais leve. O aielo DRBT04 teve tendência a associação negativa. Ao se comparar o grupo I com o grupo II, o alelo HLA-B*37 pode ser interpretado como fator de mau prognóstico. Não houve diferença estatística entre polimorfismos da região promotora do TNF-a entre pacientes e controles. Este estudo apontou uma alta frequência do genótipo TNF-a -238 G/G {OFt 3,21; Cl:1,06-9,71; p=0,04), assim como do alelo -238 G, no grupo com doença mais grave e, ao contrário, o genótipo -238 G/A com frequência maior no grupo de boa evolução. O haplótipo -238A-308G mostrou frequência reduzida conferindo um efeito protetor. Estes dados não correspondem ao reportado para as populações caucasianas, considerando que a população brasileira é miscigenada. Polimorfismos dos SNPs do TNF-a não parecem ser um fator de susceptibilidade genética mais importante do que o já conhecido HLA-Cw*06 em pacientes brasileiros, mas podem ter relação com as manifestações e evolução da doença. / Abstract: Psoriasis is an erythematous, scaly inflammatory dermatosis with a complex immunologic basis. The strongest genetic marker for psoriasis is HLA-Cw*06. Polymorphisms in the TNF-a promoter region, especially replacement of guanine with adenine in positions -238 and -308 are related to higher TNF-a production and higher risk for psoriasis in Caucasoid populations, not found in Asians. We did a case-control study of 69 patients with psoriasis type I and 70 controls, characterized clinical progression along 10-years of follow-up in mild or severe disease and determined HLA class I, II and TNF-a SNPs -238 and -308 polymorphisms to demonstrate whether these polymorphisms may be genetic risk for susceptibility to psoriasis or severity of the disease in Brazilians. Peripheral blood (10 ml) was collected. Genomic DNA from both psoriasis patients and controls was isolated using a salting out procedure. Polymorphisms were identified by PCR/SSP. Alleles and genotypes frequencies were compared by Fisher's test (GraphPad InStat 3.05 software). Two computational methods were used to determine the haplotypes of each subject, without taking into account any prior information: (1) the ELB algorithm implemented by the ARLEQUIN 3.1 software and (2) a coalescence based method as implemented by the PHASE v2 software. The haplotype frequencies were compared between group pairs by Fisher's test. Severe disease was found more frequently in male patients, associated with environmental factors and onset at adolescence. It may be suggested that alleles HLA- B*37, -Cw*06, -Cw*12 and -DRB1*07 were associated with severe disease course, while -B"57 with mild disease. No statistical difference was found between the patients and controls regarding polymorphisms frequencies in TNF SNPs. This study pointed to a higher TNF-238 G/G genotype frequency (OR 3,21; Cl:1,06-8,71; p=0,04) in the group with severe disease and -238A-308G haplotype was found in reduced frequencies revealing a protective effect. These data do not correspond to those reported for the Caucasian population, considering that Brazilian population is admixed, and this is the first consideration about TNF-a SNPs in psoriasis in this population. Polymorphisms in the TNF-a SNPs do not seem to be a more important genetic risk factor for psoriasis than the already known Cw*06 in Brazilian patients, but these markers may be related to clinical manifestations. / Doutorado / Clinica Medica / Doutor em Clínica Médica

Eine sichere Bindung zwischen Mutter und Kind in den ersten Lebensjahren ist prägend für die Entwicklung eines Kindes. Die Qualität dieser Bindung ist ein wichtiger Prädiktor für langfristige physische und psychische Gesundheit. Für den Aufbau einer starken Bindung ist die Investition von Ressourcen seitens der Fürsorgeperson auf zeitlicher, physischer und emotionaler Ebene notwendig. Multimodale biologische Hinweisreize seitens des Kindes fördern dieses Engagement. Zunächst dienen solche Signale der Identifikation des eigenen Nachwuchses (kin recognition), um nachfolgend gezielt Ressourcen zu investieren. Darüber hinaus können infantile Stimuli affektive Reaktionen vermitteln, die den Bindungsaufbau erleichtern. In diesem Zusammenhang sind auch olfaktorische Signale, z. B. Körpergerüche, wirksam, bislang gibt es jedoch nur wenig systematische Forschung zu ihrem Einfluss auf die Eltern-Kind-Beziehung. Einzelne Studien zeigen, dass Mütter ihre Kinder am Geruch erkennen können und dass kindliche Körpergerüche auch auf neuronaler Ebene positive Reaktionen vermitteln, wobei jedoch unklar ist, wie spezifisch die neuronale Aktivität für den Geruch des eigenen Kindes ist. In der vorliegenden Arbeit soll der Einfluss von kindlichen Körpergerüchen in der Mutter-Kind- Beziehung über die kindliche Entwicklungsspanne unter Berücksichtigung genetischer, hormoneller und neurobiologischer Faktoren untersucht werden. In Veröffentlichung 1 wurde geprüft, ob Mütter ihre Kinder am Geruch identifizieren können, ob sie diesen präferieren und wie beides mit genetischen und hormonellen Faktoren sowie dem kindlichen Entwicklungsstatus interagiert. Dafür wurden N = 164 Müttern mit ihren biologischen Kindern (N = 226 Kinder zwischen 0 und 18 Jahren) in die Studie eingeschlossen. Die Mütter bewerteten die Körpergerüche des eigenen und fremder Kinder, die sich im Entwicklungsstatus sowie der genetischen Ähnlichkeit unterschieden. Die genetische Ähnlichkeit wurde über das Humane Leukozytenantigen(HLA)-Profil abgebildet, der Entwicklungsstatus wurde anhand der Steroidhormonkonzentration (Testosteron, Estradiol) und einer standardisierten Einschätzung des pubertären Status erfasst. Es zeigte sich, dass die Mütter den Geruch ihres eigenen Kindes über dem Zufallsniveau identifizieren konnten und diesen Geruch präferierten. Dies galt für alle Altersgruppen, mit Ausnahme der frühen Pubertät. In diesem Alter (9-13 Jahre) konnten die Mütter den Geruch ihres Kindes weder identifizieren, noch bevorzugten sie ihn im Vergleich zu fremden Körpergerüchen. Bei den eigenen Söhnen war die Abnahme der Präferenz mit dem Anstieg des Testosteronlevels assoziiert. Mit zunehmendem Alter des Kindes (14-18 Jahre) ähnelte das Bewertungsverhalten der Mütter wieder dem vor Pubertätsbeginn, was vermuten lässt, dass die Mütter sich in diesem Zeitraum an den veränderten Geruch des Kindes gewöhnen und somit die Vertrautheit des Geruchs eine wichtige Rolle für die Wahrnehmung spielt. Zusätzlich legen die Ergebnisse nahe, dass genetische Ähnlichkeit über Körpergerüche transportiert wird: Der Geruch des eigenen Kindes wurde zwar global bevorzugt, im paarweisen Vergleich zeigte sich jedoch, dass sich die Bewertung für den Geruch des eigenen Kindes nicht signifikant von der Bewertung des gleichaltrigen und HLA-ähnlichen Kindes unterschied. Dies lässt darauf schließen, dass sich genetische Ähnlichkeit positiv auf die Geruchsbewertung im Kontext der Eltern-Kind-Bindung auswirkt. Für die zweite Veröffentlichung wurde anhand derselben Stichprobe getestet, ob Mütter den Entwicklungsstatus des Kindes anhand von Körpergerüchen klassifizieren können und welche Prädiktoren für die Klassifikation entscheidend sind. Dafür wurden sie gebeten, die jeweilige Altersgruppe des Kindes einzuschätzen, von dem der Geruch stammte. Die Ergebnisse demonstrieren, dass Mütter den kindlichen Entwicklungsstatus (prä- bzw. postpubertär) mit einer Genauigkeit von 64 % detektieren können und insgesamt dazu tendieren, kindliche Körpergerüche als präpubertär zu klassifizieren. Die mütterliche Klassifikationsleistung war besser, wenn die Probandinnen Geruchsproben aus der gleichen Altersgruppe wie der des eigenen Kindes beurteilten. Die subjektive Bewertung der Proben hinsichtlich Angenehmheit und Intensität sowie die Einschätzung des pubertären Status waren signifikante Prädiktoren für die entwicklungsbedingte Klassifikation eines Geruchs, während sich der Steroidhormonstatus des Kindes nicht auf die mütterliche Einschätzung auswirkte. Die dritte Veröffentlichung dieser Doktorarbeit diente als methodische Pilotstudie für die spezifische Untersuchung des Einflusses von Babygerüchen auf die neuronale Verarbeitung im mütterlichen Gehirn. Aus anderen Modalitäten ist bekannt, dass kindliche Stimuli Niedlichkeit vermitteln, welche mit belohnungsspezifischer neuronaler Aktivität einhergeht. Dies ist für Babygerüche bisher jedoch kaum erforscht. Die Präsentation von Körpergerüchen zur Ableitung neuronaler Korrelate im Rahmen von funktioneller Magnetresonanztomographie (fMRT) ist aufgrund von Stimuluseigenschaften sowie methodischen Schwierigkeiten herausfordernd. Bislang existieren nur wenige Studien zur neuronalen Verarbeitung von Körpergerüchen ohne einheitliche Konvention über eine geeignete Stimuluspräsentation. Im Rahmen dieser Doktorarbeit sollte daher ein effizientes Design entwickelt werden, welches neuronale Aktivität in Reaktion auf Babygerüche optimal abbildet. Dafür wurden zwei Stimuluspräsentationen verglichen, die sich in Art, Dauer und Frequenz unterschieden. Die kurze, kontinuierliche Reizdarbietung rief im Vergleich zu einer langen, gepulsten Präsentation global stärkere Aktivierungen hervor, weshalb diese als Design empfehlenswert ist, um robuste neuronale Korrelate zu erhalten. Allerdings zeigten sich differentielle Effekte in Abhängigkeit der Hirnregionen, weshalb je nach interessierendem Areal spezifisch zwischen Länge, Dauer und Art der Stimuluspräsentation abgewogen werden sollte. Das kurze Präsentationsdesign wurde im Rahmen der weiterführenden fMRT-Studie verwendet. Diese veranschaulichte, dass Babygerüche Belohnungsareale sowie Netzwerke aktivieren, die Angehmheit, Niedlichkeit und Motivation zur Fürsorge (Pleasure-Netzwerk) kodieren. Die Aktivierungsstärke des Netzwerks sagte dabei vorher, wie angenehm die Mütter den Geruch des eigenen Babys bewerteten. Im Gegensatz zu den Verhaltensdaten aus Veröffentlichung 1, in denen sich eine klare Präferenz für das eigene Kind zeigte, konnte kein Unterschied zwischen der neuronalen Reaktion auf den Geruch des eigenen im Vergleich zu einem fremden Baby gefunden werden. Daher gilt es, die Universalität des Babygeruchs als einen Stimulus, der Niedlichkeit vermittelt, in nachfolgenden Studien systematisch zu überprüfen. Zusammenfassend stellt diese Arbeit dar, dass kindliche Körpergerüche als Chemosignale in der Mutter-Kind-Beziehung wirken und sowohl zur Identifikation des eigenen Kindes beitragen als auch affektive Komponenten vermitteln. Außerdem wurde herausgefunden, dass Körpergerüche Informationen über genetische Ähnlichkeit und den Entwicklungsstatus des Kindes transportieren. Es bleibt offen, welche Faktoren auf molekularer Ebene tatsächlich die Veränderung des Körpergeruchs ausmachen. Chemosensorische Profilanalysen können in zukünftigen Untersuchungen Aufschluss darüber geben. Darüber hinaus sind Langzeitstudien notwendig, um die hier dargestellten assoziativen Zusammenhänge auch über den individuellen Entwicklungsverlauf abzubilden und somit Mechanismen der olfaktorisch vermittelten Eltern-Kind-Beziehung ableiten zu können. Langfristig sollen diese Informationen dazu beitragen, Strategien zur Förderung der Eltern-Kind-Beziehung zu generieren und bisher bestehende Interventionen (wie z. B. Neurofeedbacktraining) auf olfaktorische Stimuli auszuweiten.:Inhaltsverzeichnis Danksagung 4 1 Zusammenfassung 6 2 Summary 9 3 Einführung in die Thematik 12 4 Studienziele: Abgeleitete Forschungsfragen und Hypothesen 21 5 Methodik der Untersuchungen 23 6 Zusammenfassung der Ergebnisse 26 7 Diskussion und Ausblick 29 8 Literaturverzeichnis 40 Anhang I. Verzeichnis der wissenschaftlichen Veröffentlichungen, Konferenzbeiträge und andere Leistungen A Veröffentlichungen der Dissertation und dazugehörige Angaben B Weitere Veröffentlichungen während der Promotionsphase C Konferenzbeiträge und andere Leistungen während der Promotionsphase II. Letters of Acceptance III. Erklärung zur Eröffnung des Promotionsverfahrens IV. Bestätigung der Einhaltung der folgenden aktuellen gesetzlichen Vorgaben / A secure bond between mother and child in the first years of life is crucial for the development of a child. The quality of this bond is an important predictor of long-term physical and mental health. To create such a bond, the caregiving person has to invest resources at a temporal, physical and emotional level. Multimodal biological infantile cues facilitate this commitment. Initially, such signals serve to identify one's own offspring (kin recognition) in order to invest resources in a targeted manner. In addition, infantile stimuli can mediate affective reactions that support bonding. In this context, olfactory signals, e.g. body odors, are also effective, but so far there is little systematic research on their influence on the parent-child relationship. Individual studies show that mothers can recognize their children by their body odor and that infantile body odors also mediate positive reactions at the neural level, although it is unclear how specific they are for their own child. The present study investigates the influence of children ́s body odors in the mother-child relationship over the developmental span, integrating genetic, hormonal and neurobiological factors. Publication 1 addressed the question of whether mothers can identify their children by body odor, whether they prefer this odor and how it interacts with genetic, hormonal factors and the child's developmental status. For this purpose, N = 164 mothers with their biological children (N = 226 children between 0 and 18 years) were included in the study and evaluated the body odors of their own and unfamiliar children, which differed in their developmental stage and genetic similarity. Genetic similarity was mapped via the human leukocyte antigen (HLA) profile, the developmental status was determined on the basis of the steroid hormone concentration (testosterone, estradiol) and a standardized assessment of the pubertal status. The results showed that the mothers were able to identify their own child's odor above chance level and preferred this odor. This was true for all age groups with the exception of early puberty. At this age (9-13 years), mothers could neither identify the odor of their child nor preferred it to unfamiliar body odors. For the body odor ratings of their own sons, the decrease in preference was associated with an increase in testosterone level. In older children (14-18 years), maternal ratings resembled those before puberty suggesting that the mothers get used to the altered body odor of their child during this period and thus, the familiarity of the odor plays an important role for perception. In addition, the results demonstrated that genetic similarity is transported via body odors: Although the preference for the odor of one's own child was globally observed, pairwise comparisons showed that the ratings for the own child ́s odor did not differ significantly from the evaluation of a same-aged and HLA-similar child. This suggests that genetic similarity has a positive effect on odor assessment in the context of parent-child bonding. In the second publication, in the same sample it was examined whether mothers are able to classify the child's developmental status on the basis of body odors and which predictors are decisive for the classification. Therefore, the mothers were asked to assess the age group of the child who was the odor donor. The results revealed that mothers are able to detect the developmental status (pre- vs. postpubertal) with an accuracy of 64% and tend to classify body odors as prepubertal. The maternal classification performance was better when they rated odor samples from the same age group as their own child. The perceptual evaluation of the samples (pleasantness, intensity) as well as the assessed pubertal status predicted the development-related classification of an odor, while the child ́s steroid hormone concentration had no effect on it. The third publication of this doctoral thesis served as a methodical pilot study for the specific examination of the influence of baby odors on neural processing in the maternal brain. From other modalities, it is known that infantile stimuli transport cuteness leading to reward-related neural correlates. However, this has scarcely been investigated for baby odors so far. Body odor presentation in functional magnetic resonance imaging (fMRI) is challenging due to stimulus properties and methodological difficulties. To date, only a few studies exist on the neural processing of body odors without a uniform convention on a suitable stimulus presentation. The aim of this thesis was to develop an efficient design that optimally maps neural activity in response to baby body odors. For that reason, two stimulus presentations were compared which differed in presentation mode, duration and frequency. The short, continuous stimulus presentation revealed stronger global activations compared to a long, pulsed presentation, thus it is recommended as a design to obtain robust neuronal correlates. However, differential effects were observed depending on the brain regions, which is why the design should be specifically adapted to the regions of interest, and length, duration and type of stimulus presentation should be considered carefully. The short presentation design was used in the follow-up fMRI study. This illustrated that baby body odors activate reward areas and a network encoding cuteness and motivation to care (pleasure network). The recruitment of this network predicted how pleasantly mothers rated their own baby's odor. In contrast to the behavioral data from publication 1, which showed a clear preference for one's own child, the neural responses did not differ between one's own or an unfamiliar baby ́s odor. Therefore, the universality of baby odor as a stimulus conveying cuteness must be systematically examined in subsequent studies. In summary, this doctoral thesis reveals that children ́s body odors function as chemosignals in the mother-child relationship and mediate both the identification of the own child and affective components. In addition, it was observed that information about genetic similarity and the child's developmental status are transcribed in body odors. It remains to be explored which factors at the molecular level actually determine changes in body odor. Future investigations using chemosensory profile analyses may clarify this question. Beyond that, longitudinal studies are necessary in order to depict the associations presented here over the course of individual development and thus enabling the derivation of mechanisms of the olfactory mediated parent-child relationship. In the long term, this information should help to generate strategies for promoting the parent-child relationship and to extend existing interventions (such as neurofeedback training) to olfactory stimuli.:Inhaltsverzeichnis Danksagung 4 1 Zusammenfassung 6 2 Summary 9 3 Einführung in die Thematik 12 4 Studienziele: Abgeleitete Forschungsfragen und Hypothesen 21 5 Methodik der Untersuchungen 23 6 Zusammenfassung der Ergebnisse 26 7 Diskussion und Ausblick 29 8 Literaturverzeichnis 40 Anhang I. Verzeichnis der wissenschaftlichen Veröffentlichungen, Konferenzbeiträge und andere Leistungen A Veröffentlichungen der Dissertation und dazugehörige Angaben B Weitere Veröffentlichungen während der Promotionsphase C Konferenzbeiträge und andere Leistungen während der Promotionsphase II. Letters of Acceptance III. Erklärung zur Eröffnung des Promotionsverfahrens IV. Bestätigung der Einhaltung der folgenden aktuellen gesetzlichen Vorgaben

The thesis consists of two parts. The first part introduces the topic of human chemical communication and reviews current evidence on individual variation in human body odour and its perception. This part is framed by sexual selection theory. In the first chapter, the concept of the theory of communication is introduced followed by a discussion on the specifics of chemical communication. Next, the formation of individually specific body odour signatures with reference to skin glands, their volatile products and the subsequent metabolization by skin microflora is described. The next chapters are dedicated to selected interindividual body odour cues such as sex and kin recognition, genetic compatibility in genes of Major Histocompatibility Complex, and health and reproductive status in a mate choice context. Furthermore, interactions between perfumes and body odours are discussed. Finally, methods of body odour sampling are introduced and a rationale behind presenting individual samples or body odour blends is discussed. The second part is comprised of six scientific papers, specifically three reviews and three empirical studies. Review papers summarize factors affecting human body odour quality with emphasis on diet and affective states. The first text shows that human body odours contain cues to...

T cell receptors (TCR) bind to peptides from various sources on MHC (Major Histocompatibility Complex) molecules. A long-standing goal in the field is to understand the mechanisms of MHC-peptide exchange and MHC-TCR interactions. Here, I present work from three uniquely different systems that address the following: HLA-DR1 conformational stability, self-tolerant mechanisms of TCRs isolated from self-reactive TCR transgenic mice, and TCR cross-reactivity mechanisms between LCMV and VV. First, I present a crystal structure of HLA-DR1 in complex with A1L9 peptide, a peptide with two amino acid substitutions from the parental peptide. The singly substituted A1 peptide, which has a pocket 1 alanine substitution, decreases intrinsic half-life between MHC-peptide and increases susceptibility to HLA-DM mediated peptide exchange. This data agrees with previous models of HLA-DM-mediated peptide exchange in which the major determinant is located at the HLA-DR1 pocket 1. However, the L9 substituted peptide, which has a pocket 9 leucine substitution, displays the opposite phenotype: increased intrinsic half-life and decreased HLA-DM susceptibility. The crystal structure presented here shows that HLA-DR1 in complex with a doubly substituted peptide, A1L9, is in the same conformation as HLA-DR1 with the wild-type peptide, demonstrating that pocket 9 residues can rescue pocket 1 residue binding deficiencies and that HLA-DR1 stability is determined by amino acids along the peptide, not only at pocket 1. Next, I present crystal structures of two self-tolerant TCRs in complex with IAb-3K pMHC. To elucidate molecular mechanism for self-reactivity and self-tolerance, the TCRs J809.B5 and 14.C6 are compared to each other and its parental self-reactive TCR, YAe-62.8. In comparison to YAe-62.8, J809.B5 interacts with the same pMHC, but utilizes more peptide specific interactions, a mechanism that may distinguish self-reactive receptors from self-tolerant receptors. Additionally, the crystal structure of 14.C6 TCR, which bears a different CDR3α sequence from J809.B5, demonstrates that CDR3 sequences can modulate interactions of germline encoded CDR1 and CDR2 loops. Together, these results highlight that in addition to CDR3 VDJ recombination, diversity is generated in the mature TCR repertoire by differential chain pairing, either of which can affect the interactions of germline encoded CDR loops. Next, I present a detailed analysis of cross-reactive TCRs between Kb-GP34 and Kb-A11R. The mature LCMV-immune repertoire was analyzed by DNA deep sequencing of TCRβ CDR3 sequences, which led to the identification of new cross-reactive sequence motifs. Cross-reactive sequence motifs varied by each Vβ gene, suggesting a role of CDR1, CDR2, and CDR3 loop interplay in cross-reactivity. Lastly, I present the crystal structures of a GP34/A11R cross-reactive TCR in complex with both Kb-GP34 and Kb-A11R. Analysis of the crystal structures revealed that the two complexes are largely the same, despite differences in peptide sequences. Surprisingly, the TCR to peptide interactions were dominated by three out of eight peptide side-chains. Cross-reactivity between these two complexes is likely due to a large amount of interactions from TCR to MHC compared to interactions of TCR to peptide. We note two unique MHC-peptide interactions that may allow Kb to be an allele prone to cross-reactivity. The first is an interaction at the C-terminus of the A11R peptide which pulls A11R P7 asparagine away from TCR interactions. The second interaction is from an arginine at position 155, which sits at the interface between TCRα and TCRβ , and contributes the most buried surface area in the interaction interface. Because Kb’s arginine 155 is a long side chain that hydrogen bonds with the peptide backbone, and is also at the center of the TCR-peptide interface, GP34 and A11R peptide sequence differences may be occluded from TCR discrimination by Kb presentation. The data presented in this dissertation demonstrate that interactions between MHC-peptide and MHC-TCR act harmoniously and coopertively, whereby proximal interactions are affected by interactions elsewhere. While previous models of HLA-DR/HLA-DM interactions demonstrate the importance of interactions at HLA-DR1 pocket 1, I showed that pocket 9 also contributes to HLA-DR stability and therefore, HLA-DM susceptibility. I also showed that TCR CDR3 loop sequences affect germline CDR1/CDR2 loop interactions and vice versa. Lastly, I showed that allele specific MHC side chain interactions with the bound peptide influence TCR ligand binding and hence, TCR cross-reactivity.

Major histocompatibility complex molecules (MHCII) are cell surface glycoproteins that present extracellular antigens to CD4+ T lymphocytes and initiate adaptive immune responses. Apart from their protective role, overexpression of MHCII contributes to autoimmune disorders where the immune system attacks our own tissues. Autoimmune diseases are characterized by self-reactive responses to autoantigens, promoting tissue damage, inflammation mediated by proinflammatory cytokines, autoreactive lymphocytes, and autoantibodies. MHCII molecules are tightly regulated at the level of transcription by Class II transactivator (CIITA). CIITA associates with an enhanceosome complex at MHCII promoters and regulates the expression of MHCII. It is thus crucial to understand the regulation of CIITA expression in order to regulate MHCII in autoimmune diseases. Our lab has shown that the 19S ATPases of the 26S proteasome associate with MHCII and CIITA promoters and play important roles in gene transcription, regulate covalent modifications to histones, and are involved in the assembly of activator complexes in mammalian cells. The mechanisms by which the proteasome influences transcription remain unclear. Here, we define novel roles of the 19S ATPases Sug1, S7, and S6a in expression of CIITApIV genes. These ATPases are recruited to CIITApIV promoters and coding regions, interact with the elongation factor PTEFb, and with Ser5 phosphorylated RNA Pol II. Both the generation of CIITApIV transcripts and efficient recruitment of RNA Pol II to CIITApIV are negatively impacted by knockdown of 19S ATPases. Alternatively, inflammation is also suppressed via the Regulator of G-protein signaling 10 (RGS10) in microglial cells which express high levels of RGS10 and promote homeostasis in the central nervous system. However, chronic activation of microglial cells leads to release of cytokines which cause neuroinflammation. Our investigation of roles played by RGS10 in chronically activated microglial cells indicates that RGS10 binds to promoters of IL-1β, and TNF-α and regulates these genes, while the molecular mechanism remains to be investigated. Together, our observations indicate roles for the UPS in modulating gene expression and for RGS10 in regulating proinflammatory cytokines in microglial cells, each of which provides novel therapeutic targets to combat inflammation in autoimmune and neurodegenerative diseases.

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