Les lymphoproliférations survenant après transplantation sont une situation rare mais préoccupante car mettant en jeu la survie des patients. Ces hémopathies ont des caractéristiques épidémiologiques et physio-pathologiques qui les distinguent des lymphomes du sujet immunocompétent. Nous nous sommes intéressé à l’analyse des facteurs de risque associés aux lymphomes post-greffe et à la recherche de facteurs pronostiques de survie des patients par l’analyse détaillée des registres américain et français de patients transplantés rénaux. Le Registre Français des lymphomes survenant après transplantation rénale a permis de recenser tous les cas de syndromes lymphoprolifératifs se développant chez des patients adultes survenant entre le 1er janvier 1998 et le 31 décembre 2007. Tous les centres français de Transplantation rénale ont participé. Nous avons recensé 500 cas de lymphomes sur une période de 10 ans. Les différentes analyses de cette base de données ont donné lieu à la publication d’une analyse intermédiaire sur les 230 premiers cas, à une publication consacrée à l’incidence et aux facteurs de risque de développement des lymphomes surla cohorte complète et à une publication concernant les facteurs pronostiques de décès des patients porteurs d’une lymphoprolifération avec établissement d’un score de risque spécifique de cette population. D’autre part, cette base de données unique au monde représente un support intéressant pour le développement de travaux de recherche: étude de l’origine des cellules tumorales, étude des facteurs de susceptibilité pharmacogénétique au développement des lymphoprolifération post-greffe, étude des microRNA de l’EBV dans les lymphomes post-greffe. / Post transplant lymphoproliferative disorders (PTLD) are a rare but serious complication occurring after kidney transplantation. Features of PTLD are specific and different of those of immunocompetent patients. We studied incidence, risk factors for development of PTLD and prognostic factors of patients with PTLD using two databases: American and French. The French Registry of PTLD enrolled all adult patients with PTLD occurring between the 1st January 1998 and the 31st December 2007 from all transplant centers in France. Five hundred patients were included in the Registry. This enables us to analyse first the incidence and risk factors of PTLD and second the risk factors of kidney recipients’ survival. We constructed a new prognostic score specific of transplant patients. Finally, the French Registry gave us the opportunity to support others research projects: determination of the origin of tumoral cells, analysis of pharmacogenetic factors associated with the risk of developing PTLD, study of EBV microRNA in lymphoproliferations.

Aims: To explore the clinical significance of EBV infection in adult renal transplant recipients when detected in the late post-transplant period. Methods: (1) A prospective observational study recruiting 499 stable adult kidney transplant recipients with serial blood sampling for EBV DNAemia and assessment of clinical outcomes and associated factors. (2) A retrospective analysis of PTLD incidence, timing and outcomes in relation to EBV infection. Results: EBV DNAemia in stable kidney transplant recipients is common, found in 46% of recruited individuals screened over 1 year, with persistent DNAemia seen in 10%. DNAemia prevalence increased significantly with time from transplant (p<0.0001) from 16% within 1 year of transplant to 66% in those transplanted for 20-24 years. High baseline DNA levels predicted persistence of DNAemia. Time adjusted analyses showed significant association of DNAemia with EBV seronegative status and previous PTLD and low DNAemia rates with Mycophenolate Mofetil (MMF) use and lymphopenia. The mechanism did not appear to be directly linked to MMF induced B cell depletion. Chronic high viral load detection was significantly associated with time from transplant, EBV seronegative status at transplant, ciclosporin use and plasma detection of DNA. No significant differences in overall patient survival at 3 years, clinical symptoms or clinical findings such as anaemia, thrombocytopenia or rate of decline in renal function were seen between stable transplant recipients with and without EBV DNAemia. PTLD incidence also increases with time from transplant and was greatest during the 10th-14th post-transplant years. Disease was EBV positive in 68% cases. No statistically significant differences in overall patient survival, or overall disease complete response rates were seen in relation to recipient EBV serostatus or EBV status of PTLD histology. Conclusions: EBV DNAemia prevalence increases with time from transplant but was not associated with worse patient or graft survival or specific symptoms. PTLD incidence including EBV negative disease also increases with time from transplant but response rates and survival were not influenced by EBV serostatus or histological status.

Experiments were performed to assess the suitability of Rhabditis pellio, a nematode found in earthworms, as a challenge organism for use in development of a biomarker assay to determine the potential of chemicals to suppress the immunocompetence of the non-specific immune system. To accomplish this goal, information on the life cycle of R. pellio was determined; including effects of incubation time and temperature on growth rates; along with information on the immune response elicited in the earthworm, Lumbricus terrestris. Immune parameters measured were coelomocyte migration toward and attachment to R. pellio larvae and adults. Preliminary background information showed that R. pellio has potential as a challenge organism for development of a biomarker assay.

Orientador: Mary Anne Heidi Dolder / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-26T11:12:55Z (GMT). No. of bitstreams: 1 Freitas_KarineMoura_D.pdf: 5043196 bytes, checksum: 36715a7f1ea621d20e0985a697cef1e4 (MD5) Previous issue date: 2014 / Resumo: A Ciclosporina A (CsA) é uma droga imunossupressora amplamente utiliza na terapia pós-transplante de órgãos e no tratamento de doenças autoimunes. Apesar de sua ampla utilização e eficiência como imunossupressora, a CsA causa diversos efeitos colaterais, dentre eles nefrotoxicidade, hepatotoxicidade e danos a órgãos do aparelho reprodutor masculino (testículo e próstata). Estes efeitos provavelmente estão relacionados à atividade pró-oxidante do medicamento, que causa um desequilíbrio entre produção de espécies reativas de oxigênio e produção de enzimas antioxidantes. Heteropterys tomentosa, planta natural do cerrado brasileiro, é utilizada popularmente utilizada como tônico, estimulante das funções cerebrais e afrodisíacas. A eficiência da infusão de H. tomentosa para atenuar os danos causados pela CsA no testículo e próstata ventral de ratos já foi comprovada cientificamente. Além disso, H. tomentosa também parece ser eficiente contra a hepatotoxicidade induzida por esta droga, analisando-se parâmetros bioquímicos plasmáticos. O objetivo deste trabalho foi verificar os efeitos da Ciclosporina A no fígado, rim, timo, testículo, epidídimo e próstata ventral. Caso haja efeitos, determinar se a infusão de H. tomentosa altera tais efeitos. Além disso, objetivamos analisar se o tratamento com a infusão de H. tomentosa interfere na capacidade imunossupressora da Ciclosporina A. O tratamento com H. tomentosa durante 21 dias causou sutis alterações hepáticas nos animais, resultados não encontrados após a administração do tratamento com a planta durante 56 dias. O tratamento simultâneo com CsA e H. tomentosa causou alterações hepáticas diferentes daquelas dos animais tratados somente com CsA. A nefrotoxicidade induzida por CsA foi tempo dependente. O tratamento com CsA durante 21 dias causou o aparecimento de pequenos acúmulos de substâncias no lúmen dos túbulos da região medular, entretanto não foram observadas graves alterações nos túbulos proximais. Após 56 dias de tratamento foram observados acúmulo de substâncias no lúmen de túbulos da região medular do rim. Além disso, foram observados pontos de vacuolização citoplasmática nos túbulos proximais em regiões específicas. Estes efeitos foram observados no grupo tratado somente com CsA e no grupo tratado simultaneamente com CsA e H. tomentosa. O tratamento somente com a infusão de H. tomentosa não causou danos ao rim. O tratamento com CsA e/ou H. tomentosa não causou efeitos aos testículos, entretanto, após 56 dias de tratamento formam observadas alterações ao epidídimo e próstata ventral, resultado não observado no grupo CsA+Ht. A contagem de células no sangue periférico evidenciou a imunossupressão causada pela Ciclosporina A administrada por 21 ou 56 dias. Mesmo após o tratamento conjunto com a infusão e a droga (21 e 56 dias) a imunossupressão foi evidenciada. Além disso, a análise do tecido tímico revelou alterações estruturais causadas pela CsA administrada isoladamente ou em conjunto com a infusão. Estes resultados revelaram que os efeitos colaterais associados ao tratamento com CsA ao rim, epidídimo e próstata ventral são tempo-dependentes. Além disso, foi observado que o tratamento com H. tomentosa foi capaz de reduzir as alterações causadas pela CsA ao epidídimo e próstata ventral. Este estudo revela ainda, que a infusão de H. tomentosa não altera a capacidade imunossupressora da Ciclosporina A / Abstract: Cyclosporin A (CsA) is an immunosuppressive drug used after organ transplantation and against auto immune diseases. Despite CsA widely use as immunosuppressant, this drug causes diverse side effects as: nephrotoxicity, hepatotoxicity and male reproductive organs (testis and ventral prostate) impairment. Those CsA-related side effects are probably related to the CsA pro oxidant properties that causes imbalance among reactive oxygen species generation and antioxidant enzymes production. Heteropterys tomentosa, native plant from Brazilian cerrado, a savanna-like biome, is popularly used as tonic and stimulant of brain functions and as an aphrodisiac plant. H. tomentosa¿s efficiency against CsA side effects to the testis and ventral prostate has been scientifically proved. Moreover, H. tomentosa either reduced the alterations caused by CsA to plasma biochemical parameters that are hepatotoxicity markers. The aim of the present study was evaluate the effects of CsA to the liver, kidney, testis, epididymis, ventral prostate and thymus. If there are alterations CsA-induced, evaluate if H. tomentosa infusion alter these effects. Lastly we aimed to evaluate if H. tomentosa infusion interfere on CsA immunosuppressive capacity. CsA treatment for 21 days caused minor alterations to the liver tissue; this result was not observed after the administration of the infusion for 56 days. The simultaneous treatment with CsA and H. tomentosa caused hepatic alterations different from those observed in the animals treated with only CsA. The nephrotoxicity CsA-induce was time related; the kidney of animals treated with CsA had small substances accumulation (casts) on juxtamedullary tubules lumina, alteration to the proximal tubules were not evident. After 56 days of treatment there was huge casts on juxtamedullary tubules and some proximal tubules had vacuolization. These kidney¿s alterations were present in the groups treated with CsA and CsA+H. tomentosa. H. tomentosa administration did not caused alteration to the kidney tissue. The treatment with CsA, H. tomentosa and both simultaneously did not caused effects to the testis, however, CsA administration for 56 days caused alterations to the epididymis and ventral prostate; these alteration were not observe after the treatment with CsA+H. tomentosa. CsA caused reduction on the number of lymphocytes in the peripheral blood after the administration for 21 or 56 days, simultaneously with H. tomentosa or not. Furthermore, the thymus tissue had structural alterations caused by CsA, administrated alone or simultaneously to H. tomentosa (for 21 or 56 days). These results confirm CsA immunosuppression even after the administration with H. tomentosa simultaneously. The present study shows that CsA-induced effects to the kidney, epididymis and ventral prostate are time dependents. Moreover, the treatment with H. tomentosa reduced the alteration caused by CsA to the epididymis and ventral prostate. Finally, this study proved that H. tomentosa did not alter the immunosuppressant properties of Cyclosporin A / Doutorado / Biologia Celular / Doutora em Biologia Celular e Estrutural

O mecanismo de imunorregulação gerado pelo catabolismo do triptofano pela IDO protege o feto contra a resposta imunológica materna. Neste estudo, esse mecanismo foi em avaliado em gestantes imunossuprimidas e portadoras de transplante renal. Examinou-se a expressão da IDO e sua atividade nos compartimentos placentários de gestantes saudáveis e portadoras de transplante. Células produtoras de IDO e IFN-<font face=\"Symbol\">g foram imunolocalizadas na região vilosa e região decidual em ambos os grupos analisados, com mudanças no tipo celular envolvido nestas expressões nas gestantes transplantadas. Os níveis de IDO e sua atividade, assim como seus fatores de regulação NF-kB, IFN-<font face=\"Symbol\">g e IL-10 estavam diminuídos na região vilosa. No compartimento decidual a atividade enzimática da IDO estava aumentada nas gestantes transplantadas, mas não dos seus reguladores. Sendo assim, eixo de imunorregulação gerado por IDO-IFN-<font face=\"Symbol\">g na interface placentária de gestantes portadoras de transplante renal responde diferencialmente a insultos ocasionados pela utilização de imunossupressores durante a gestação. / The mechanism of immunoregulation generated by the catabolism of tryptophan by IDO protects the fetus against maternal immune response. In this study, this mechanism was evaluated in renal transplanted pregnant women and immunosuppressed. We examined the expression and activity of IDO in placental compartments of healthy pregnant women and patients with transplants. IDO and IFN-<font face=\"Symbol\">g producing cells were imunolocalizated in villous and decidual region in both groups analyzed, with changes in cell type involved in these expressions in pregnant patients transplanted. The levels and activity of IDO, as well as their regulatory factors NF-kB, IFN-<font face=\"Symbol\">g and IL-10 were decreased in the villous region. In the decidual compartment IDO activity was increased in pregnant women transplanted, but not their regulators. Thus, the axis of immunoregulation generated by IDO and IFN-<font face=\"Symbol\">g in placental interface of pregnant women with renal transplantation responds differently to insults caused by the use of immunosuppressive drugs during pregnancy.

Submitted by Adriana Martinez (amartinez@unoeste.br) on 2017-08-15T19:12:52Z No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Bianca.pdf: 748434 bytes, checksum: 25858a86b136011052140d31f626de30 (MD5) / Made available in DSpace on 2017-08-15T19:12:52Z (GMT). No. of bitstreams: 2 license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Bianca.pdf: 748434 bytes, checksum: 25858a86b136011052140d31f626de30 (MD5) Previous issue date: 2016-11-29 / The purpose of this study was to evaluate the effect of supplementation with L-arginine, in different doses, on body weight, hemogram, production of IgA, IgG and IgM immunoglobulins and in the small bowel inflammatory process of Wistar rats subjected to chemotherapy. A total of 32 Rattus novergicus Wistar rats were randomly distributed in 4 groups (8 rats / group) with similar body weight: control group (CG), the rats were given ration and water; G5-FU was given ration, water and a dose of 5-FU; Garg295 + 5-FU and Garg458 + 5-FU were given, respectively, 295 mg and 458 mg of L-arginine, which was added to the water and a dose of 5-FU. 5-FU was applied after the adaptation period (day 7) in a single dose of 200 mg of 5-FU / kg body weight, intraperitoneally. The rats were weighed individually, always in the morning and without previous fasting, to determine the weight gain before and after the application of 5-FU. Four days after application of 5-FU, the rats were anesthetized with thionembutal, blood and small intestine samples were collected. Supplementation with 295 mg of L-arginine was shown to be beneficial in reducing body weight loss during treatment with the 5-FU chemotherapeutic. The concentration of platelets was lower (P <0.05) in the G5-FU and GArg458 + 5-FU groups, but similar between the GC and GArg295 + 5-FU groups. The total leukocyte concentration showed a significant reduction in the G5-FU, GArg295 + 5-FU and GArg458 + 5-FU groups when compared to the GC group, however, there was a significant reduction of neutrophils only in the G5-FU group. There was an increase (P <0.05) in fibrinogen concentration in the groups treated with L-arginine (GArg295 + 5-FU and GArg458 + 5-FU). In the GArg295 + 5-FU and GArg458 + 5FU groups, the serum IgA concentration increased significantly in relation to the G5-FU group. Rats supplemented with L-arginine had a minor inflammatory process in the small intestine. It was concluded that supplementation with 295 mg of L-arginine / day attenuated the immediate body weight loss of Wistar rats subjected to 5-FU chemotherapy and that daily supplementation with both 295 mg and 458 mg of L-arginine may ease some side effects of 5-FU such as thrombocytopenia, neutropenia and immunomodular production of IgA, in addition to reducing intestinal inflammatory processes. / Este estudo teve o objetivo deste estudo foi avaliar o efeito da suplementação com L-arginina, em diferentes doses, no peso corporal, no hemograma, na produção das imunoglobulinas IgA, IgG e IgM e no processo inflamatório do intestino delgado de ratos Wistar submetidos à quimioterapia. Foram utilizados 32 Rattus novergicus da linhagem Wistar, distribuídos aleatoriamente em 4 grupos (8 ratos/grupo) com peso corporal semelhante: grupo controle (GC), os ratos receberam ração e água; G5-FU receberam ração, água e uma dose de 5-FU; Garg295+5-FU e Garg458+5-FU que receberam ração, 295 mg e 458 mg de L-arginina, respectivamente, adicionada na água e uma dose de 5-FU. A 5-FU foi aplicada após o período de adaptação (dia 7) na dose única de 200 mg de 5-FU/Kg de peso corporal, por via intraperitoneal. Os ratos foram pesados individualmente, sempre no período da manhã e sem jejum prévio, para determinação do ganho de peso antes e depois da aplicação da 5-FU. Quatro dias após da aplicação da 5-FU, os ratos foram anestesiados com tionembutal e colheu-se amostras de sangue e do intestino delgado. A suplementação com 295 mg de L-arginina mostrou-se benéfica na redução de perda de peso corporal durante o tratamento com o quimioterápico 5-FU. A concentração de plaquetas foi menor (P < 0,05) nos grupos G5-FU e GArg458+5-FU, porém, semelhantes entre os grupos GC e GArg295+5-FU. A concentração de leucócitos totais apresentou redução significativa nos grupos G5-FU, GArg295+5-FU e GArg458+5-FU quando comparados ao grupo GC, entretanto, houve redução significativa de neutrófilos somente no grupo G5-FU. Ocorreu um aumento (P < 0,05) na concentração de fibrinogênio nos grupos tratados com L-arginina (GArg295+5-FU e GArg458+5-FU). Nos grupos GArg295+5-FU e GArg458+5FU, a concentração sérica de IgA aumentou significativamente em relação ao grupo G5-FU. Os ratos suplementados com L-arginina apresentaram menor processo inflamatório no intestino delgado. Conclui-se que a suplementação com 295 mg de L-arginina/dia atenuou a perda de peso corporal imediata dos ratos Wistar submetidos à quimioterapia com 5-FU e que a suplementação diária tanto com 295 mg como 458 mg de L-arginina pode amenizar alguns efeitos colaterais da 5-FU como a plaquetopenia, a neutropenia e imunomodular a produção de IgA, além de reduzir processos inflamatórios intestinais.

Myeloid-derived suppressor cells (MDSC) expand during sepsis, suppress both innate and adaptive immunity, and promote chronic immunosuppression, which characterizes the late/chronic phase of sepsis. We previously reported that the transcription factors Stat3 and C/EBPb synergize to induces the expression of microRNA (miR)-21 and miR-181b to promote MDSC expansion in a mouse model of polymicrobial sepsis that progresses from an early/acute proinflammatory phase to a late/chronic immunosuppressive stage. We also showed that Gr1+CD11b+ cells, the precursors of MDSCs, from mice genetically deficient in the inflammatory protein S100A9 lack miR-21 or miR-181b in late sepsis, and are not immunosuppressive. In the present study, we show that S100A9 induces miR-21 and miR-181b during the late sepsis phase. We find that S100A9 associates with and stabilizes the Stat3-C/EBPb protein complex that activates the miRNA promoters. Reconstituting Gr1+CD11b+ cells from the S100A9 knockout mice with late sepsis with S100A9 protein restores the Stat3-C/EBPb protein complex and miRNA expressions, and switches the Gr1+CD11b+ cells into the immunosuppressive, MDSC phenotype. Importantly, we find that this process requires IL-10 mediated signaling, which induces S100A9 translocation from the cytosol to the nucleus. These results demonstrate that S100A9 promotes MDSC expansion and immunosuppression in late/chronic sepsis by inducing the expression of miR-21 and miR-181b.

abstract: Prior to the first successful allogeneic organ transplantation in 1954, virtually every attempt at transplanting organs in humans had resulted in death, and understanding the role of the immune mechanisms that induced graft rejection served as one of the biggest obstacles impeding its success. While the eventual achievement of organ transplantation is touted as one of the most important success stories in modern medicine, there still remains a physiological need for immunosuppression in order to make organ transplantation work. One such solution in the field of experimental regenerative medicine is interspecies blastocyst complementation, a means of growing patient-specific human organs within animals. To address the progression of immune-related constraints on organ transplantation, the first part of this thesis contains a historical analysis tracing early transplant motivations and the events that led to the discoveries broadly related to tolerance, rejection, and compatibility. Despite the advancement of those concepts over time, this early history shows that immunosuppression was one of the earliest limiting barriers to successful organ transplantation, and remains one of the most significant technical challenges. Then, the second part of this thesis determines the extent at which interspecies blastocyst complementation could satisfy modern technical limitations of organ transplantation. Demonstrated in 2010, this process involves using human progenitor cells derived from induced pluripotent stem cells (iPSCs) to manipulate an animal blastocyst genetically modified to lack one or more functional genes responsible for the development of the intended organ. Instead of directly modulating the immune response, the use of iPSCs with interspecies blastocyst complementation could theoretically eliminate the need for immunosuppression entirely based on the establishment of tolerance and elimination of rejection, while also satisfying the logistical demands imposed by the national organ shortage. Although the technology will require some further refinement, it remains a promising solution to eliminate the requirement of immunosuppression after an organ transplant. / Dissertation/Thesis / Masters Thesis Biology 2020

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22694号 / 医博第4638号 / 新制||医||1045(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 生田 宏一, 教授 竹内 理, 教授 濵﨑 洋子 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Introduction: Addition of a second drug used to be a strategy to achieve clinical stabilization of multiple sclerosis in many patients with on-going activity despite monotherapy. Modern immunosuppressive drugs used in monotherapy exert more specific mode of action. Methods: This retrospective observational study evaluated 5-year data from 85 patients with active multiple sclerosis despite monotherapy with either interferon beta or azathioprine, who received add-on azathioprine or interferon beta, respectively. In a subgroup of 23 patients 10- year data were analysed. In a second part of the study, a group of 126 patients switched either from interferon beta or glatiramer acetate to fingolimod was followed-up for one after the change of their treatment and a in a subgroup of 53 patients the 2-year data were assessed. Clinical (relapse frequency, disability) parameters were compared preceding and following the addition of second drug or the switch of treatment. Laboratory results and potential serious adverse events were evaluated in a group of patients with combination therapy. Results: The add-on treatment triggered a drop in annualised relapse rate by approximately 1.5 points sustained over 5 and 10 years. No effect on disability was observed. Simultaneously, white blood cell and lymphocyte counts...