Introdução: O fragmento C4d e os anticorpos anti-HLA doador específicos (DSA) são marcadores de resposta humoral em enxertos renais com rejeição, mas o papel das células B nesse processo ainda não é claro. Neste estudo foi avaliada a correlação entre C4d, DSA e células B de enxertos com disfunção e sua associação com aspectos morfológicos, função e sobrevida do rim transplantado. Material e Métodos: A marcação para C4d, células B CD20+ e plasmócitos CD138+ foi realizada por imunoperoxidase em biópsias por indicação de 110 receptores de transplante renal. Positividade para CD20 e CD138 foi definida por curva ROC (≥5 céls./mm2). O soro coletado concomitante a biópsia foi testado para DSA classe I e classe II. Estes marcadores foram correlacionados com dados clínicos e do transplante, a histopatologia de Banff e a evolução do rim transplantado. Resultados: Depósitos de C4d e DSA circulantes foram detectados em 100% e 70% dos pacientes com rejeição mediada por anticorpos (RMA) respectivamente, e nos casos de rejeição aguda celular (RAC) em 42% (p<0,001, vs. RMA) e 28% (p=0,003, vs. RMA). Estes dois marcadores correlacionaram-se positivamente (r=0,31, p=0,016). Houve correlação significativa entre DSA e plasmócitos CD138+ (r=0.32 p=0,006), mas as células CD20 e CD138 não se correlacionaram entre si. As células CD138+ predominaram na RMA, associadas com maior painel pré-transplante e DSA, mas não a C4d, e as células CD20+ predominaram na RAC e nas biópsias com fibrose intersticial/atrofia tubular, associadas a maior incompatibilidade HLA e a retransplantes. Pacientes com C4d+ tiveram pior função e sobrevida do enxerto em três anos de transplante, e aqueles com DSA+ uma pior 7 sobrevida do enxerto. Positividade para CD20 ou CD138 não foi preditiva da função ou sobrevida do enxerto. Na análise multivariada, somente o C4d foi fator de risco para perda do enxerto. Conclusões: Esses resultados confirmam o valor prognóstico do C4d e dos DSA para uma pior evolução do enxerto renal, e sugerem uma associação das células B CD20+ com parâmetros de rejeição celular e dos plasmócitos CD138+ com marcadores de resposta humoral. Entretanto, nesse estudo o infiltrado de células B na biópsia do enxerto não foi preditivo de uma pior evolução do rim transplantado. / Introduction: The fragment C4d and the donor specific anti-HLA antibodies (DSA) are markers of the humoral response in rejecting kidney grafts, but the role of B cells in this process is still unclear. In this study we evaluated the correlation between C4d, DSA and B cells in dysfunctional grafts, and their association with morphological features, and graft function and survival. Material and Methods: The staining for C4d, CD20+ B cells and CD138+ plasmocytes were done by immunoperoxidase in 110 kidney graft biopsies for cause. Positivity for CD20 and CD138 were established by ROC curve (≥5 cells/mm2). Serum collected at biopsy were tested for anti-HLA class I and II antibodies. These markers were correlated with clinical and transplant characteristics, Banff histopathology and graft outcomes. Results: C4d deposits and circulating DSA were detected in 100% and 70% of the patients with antibody-mediated rejection (AMR) respectively, and in cases with acute cellular rejection (ACR) in 42% (p<0.001, vs. AMR) and 28% (p=0.003, vs. AMR), respectively. Both markers were positively correlated (r=0.31, p=0.016), and there was also a significant correlation between DSA and plasmocytes CD138+ (r=0.32 p=0.006). CD20 and C138 cells were not siginificantly correlated. Plasmocytes CD138+ predominated in AMR, and were associated with higher pre transplant PRA and DSA positivity, but not with C4d. CD20+ B cells were highly expressed in ACR and in biopsies with interstitial fibrosis and tubular atrophy, in association with more HLA mismatches and re-transplants. Patients with C4d had poorer graft function and survival, and those 9 with DSA + also had a worse graft survival in three years of transplant. CD20 or CD138 cells were not predictive of graft outcomes. In multivariated analysis, only C4d remained a risk factor for graft loss. Conclusion: These results confirm the prognostic value of C4d and circulating DSA for a worse kidney graft outcome, and suggest an association of CD20+ B cells with parameters of cellular rejection whereas CD138+ plasmocytes correlated with markers of the humoral response. However, in this study the B cell infiltrate in graft biopsy was not predictive of adverse outcomes to the transplanted kidney.

The incidence of human papillomavirus (HPV) -associated oropharyngeal tumors is steadily increasing therefore, information about the prevalence of oral HPV and its risk factors is very important for future screening and early diagnosis of the disease. This thesis addresses three topics. The first topic is to evaluate the prevalence of oral HPV in a healthy population and to investigate risk factors for oral HPV infection, given that these data are almost completely absent in Central Europe. A statistically significantly higher rate of positivity (8.8%) of oral HPV infection was found in the group of older unvaccinated probands than in younger partially vaccinated volunteers (2.0%). The seropositivity rate of anamnestic HPV antibodies was comparable in both groups. None of the analyzed risk factors were significantly associated with oral HPV positivity. The second topic of the thesis is the dynamics of HPV specific antibodies in patients with head and neck cancer and their prognostic significance. In patients with cervical cancer, a decrease in HPV E6 / E7-specific antibodies is associated with a better prognosis. Another goal of the dissertation was to assess the importance of anamnestic antibodies and antibodies against oncoproteins E6 and E7 in long-term follow-up 2-14 years after the end of...

Kidney transplantation is the most appropriate treatment for end-stage kidney failure. The risk of graft failure in retransplanted patients is generally higher than in first-transplant patients due to immunological and non-immunological reasons. An important risk factor to consider for retransplant patients is their sensitization, i.e. the presence of antibodies directed to HLA antigens of previous donor(s). For that reason, a project called Forbidden (Non-acceptable) Antigens was launched by IKEM with the aim of reducing the incidence of acute cellular and antibody-mediated rejection in retransplant patients. Work on the project was carried out between the years 2011-2013. Forbidden antigens were defined as mismatched HLA antigens of previous kidney donor(s) against which patients waiting for retransplantation produced antibodies. The aim of this diploma thesis is to evaluate whether the incidence of rejection is lower in patients with forbidden HLA antigens in comparison with a control cohort, where no forbidden antigens are defined. 234 patients (162 males and 72 females) were included in the study. Almost all tested patients were producing HLA antibodies (90.2%) and forbidden antigens were determined in 71.4% of patients. In a control group of 267 patients waiting for their first transplantation, the...

La transplantation rénale est la stratégie de suppléance rénale la plus performante. Le renforcement des thérapeutiques ciblant la réponse cellulaire T (i) a conduit à réévaluer la réponse allogénique humorale et (ii) a souligné deux complications majeures de la pression immunosuppressive : l’infection à cytomégalovirus CMV et le risque de cancer. Dans le travail présenté ici, nous analysons d’abord l’impact histologique de deux de ces facteurs de détérioration de l’allogreffon rénal : l’infection à CMV avant une biopsie sur indication et la pathogénicité des anticorps anti-HLA dirigés contre le donneur, détectés par des techniques d’identification en Single Antigen in situ dans le greffon. Nous montrons ensuite comment les lymphocytes T (LT) γδ Vd2neg font le lien entre CMV et DSA : induits par le CMV, les LT γδ Vd2neg participent aux lésions médiées par les DSA par leur capacité à réaliser une lyse dépendante de l’anticorps (ADCC) impliquant le CD16. En plus de cette nouvelle fonction allogénique indirecte, les LT γδ Vd2neg possèdent une double réactivité anti-CMV et anti-tumoral. Nous présentons ici un modèle où les lymphocytes T γδ Vδ2neg s’activent spécifiquement de façon TCR dépendante par la reconnaissance d’un marqueur d’intégrité épithéliale (EphA2) : ils détournent le mécanisme d’interaction classique d’EphA2 avec ses ligands naturels éphrines A1 et A4 pour s’en faire un signal de costimulation, en s’appuyant sur le contexte de stress pour renforcer son activation. Collectivement, nos résultats contribuent à mieux préciser la bioréactivité et le rôle des LT γδ Vδ2neg en transplantation rénale. Nos données suggèrent que chez l'homme, a fortiori lorsqu’il est immunodéprimé, les LT γδ constituent un compartiment de surveillance lymphoide du stress, capable de censurer la dérégulation des cellules infectées ou transformées et de prendre part à la réponse allogénique par un mécanisme d’ADCC. / Kidney transplant is the most performant strategy for renal replacement therapy. Increasing treatment targetting T cell response has led (i) to reappraise the importance of humoral allogenic response, (ii) to underline two main complications subsequent to immunosuppressive pressure : cytomegalovirus infection and tumorigenesis. Here, we first report the pathological impact of two of these factors on kidney allograft deterioration : CMV infection prior a biopsy for cause and Donor Specific Alloantibodies (DSA) detected within the graft with single antigen flow bead assay. Then we showed that CMV-induced Vδ2neg γδ T cells are a new player and a potentially useful clinical biomarker in antibody-mediated lesions of kidney transplants. By engaging DSA on their Fcγ-receptor CD16, γδ T cells participate in allograft lesions mediated by DSA through antibody-dependent cell cytotoxicity (ADCC). In addition to this new indirect allogenic function, CMV-induced Vδ2neg γδ T cells displayed a dual anti-CMV and anti-tumor reactivity. Finally, we here identified EphA2 as a new stress-regulated antigen targetting by a non Vδ2neg γδ T cell clone, which recognition implies the hijacking of the natural EphA2-ephrin interactions to activation. These data suggest that in humans, a fortiori when immunosuppressed, γδ T cells compose a lymphoid stress-surveillance compartment, capable of recognizing the dysregulated state of infected or transformed cells and to take part to allogenic response through an ADCC mechanism.