O papel das células T reguladoras dos ligandos BAFF e APRIL e receptores Baff-R e TACI no cancro e autoimunidade

Trabalho final de mestrado integrado em Medicina (Biologia Aplicada/Medicina Interna), apresentado á Faculdade de Medicina da Universidade de CoimbraIntrodução: Atualmente, verifica-se um aumento de incidência tanto de doenças oncológicas como de doenças auto-imunes na população em geral. Há diversos mecanismos celulares e vias de sinalização celular envolvidos na doença autoimune e oncológica, no entanto o NF-κB, os ligandos BAFF e APRIL e seus recetores, BAFF-R e TACI, e as células reguladoras T reguladoras são os mais preponderantes. Objetivos: Com este trabalho pretende-se identificar e quantificar o NF-kB, os ligandos BAFF e APRIL e seus recetores, BAFF-R e TACI, e as células T reguladoras, em doentes com doenças auto-imunes e/ou neoplásicas, de modo a contribuir para clarificar o seu papel nestas patologias. Métodos: Foram quantificadas as células T reguladoras (Treg) e avaliada a expressão do BAFF, BAFF-R, TACI e APRIL em doentes com doenças auto-imunes e/ou neoplásicas de modo a analisar o seu papel na fisiopatogenia destas doenças. O estudo foi efetuado em Sangue Periférico (SP) de 11 controlos saudáveis (CTL) e de 76 doentes, dos quais 21 com doenças auto-imunes (DAI), 47 com neoplasias (NEO) e 8 com ambas as patologias (DAI e NEO). Após a colheita do SP, procedeu-se à análise das diferentes populações linfocitárias, com identificação das células Treg, e avaliação da expressão do BAFF, BAFF-R, TACI e APRIL por citometria de fluxo, utilizando anticorpos monoclonais marcados com sondas fluorescentes. Os resultados obtidos foram estatisticamente analisados pelo teste Anova, T-student e análise multivariada (p<0.05). Resultados: Na análise dos resultados do linfograma, os linfócitos totais dos doentes DAI e dos doentes NEO encontravam-se diminuídos em comparação com os controlos. Observou-se diminuição significativa de linfócitos B nos doentes com DAI e doença auto-imune e neoplásica (DAI+NEO) e aumento significativa dos linfócitos T dos doentes com DAI comparativamente aos controlos e aos doentes com neoplasia. As células nTreg dos indivíduos saudáveis (controlos) representam um pequeno nicho de células, no entanto, os diferentes grupos de doentes apresentam aumento significativo desta percentagem de células. Os doentes com neoplasia apresentam aumento da percentagem de células a expressar BAFF comparativamente aos controlos, contrariamente, os doentes com DAI+NEO e os doentes com DAI apresentam menor percentagem destas células. O recetor de superfície BAFF-R revelou maior percentagem de expressão nas células de doentes com DAI+NEO do que nas células dos controlos. A análise do ligando TACI revelou que o grupo de doentes com neoplasia se destacava dos restantes grupos de doentes devido à elevada percentagem de expressão. Por fim, a análise do ligando APRIL revelou que os grupos de doentes com DAI+NEO e os NEO apresentam cerca de 3 vezes mais células com expressão desta molécula comparativamente aos controlos. Discussão: Nos doentes analisados verificou-se aumento significativo das células nTreg nos doentes com DAI, com doença neoplásica e principalmente nos doentes com DAI e neoplasia, possivelmente uma forma das patologias oncológicas gerarem um ambiente imunossupressor e uma tentativa do sistema imune dos doentes com DAI de suprimir a resposta inflamatória anómala. Considerando os ligandos BAFF, BAFF-R, TACI e APRIL, o grupo de doentes com neoplasia, revelou aumento substancial de expressão destes ligandos, revelando que podem ser mecanismos adquiridos de sobrevivência celular em doenças neoplásicas. Conclusão: Comprovou-se que há distúrbios relevantes a nível dos linfócitos B e T, com alteração das suas populações em ambas as patologias. Além disso, os nossos resultados sugerem que as células T reguladoras e os ligandos e recetores relacionados com o NF-kB, BAFF, APRIL e BAFF-R e TACI, respetivamente, podem participar na fisiopatologia das doenças auto-imunes e neoplásicas, podendo eventualmente constituir novos alvos terapêuticos para o tratamento destas patologiasIntroduction: Now a day, it’s verified that there is an increase of incidence in both oncologic and autoimmune diseases on general population. There are several cellular mechanisms and cellular pathways involved on autoimmune and oncologic pathologies, however, the NF-κB, the ligands BAFF and APRIL and their receptors BAFF-R and TACI and the T regulatory cells are the most preponderant. Objectives: With this work we want to identify and quantify the NF-κB, the ligands BAFF and APRIL, their receptors BAFF-R and TACI, and the T regulatory cells on patients with autoimmune and/or oncologic diseases, so that we can contribute to clarify their role on these pathologies. Methods: The T regulatory cells (Treg) were quantified and it was evaluated the expression of BAFF, BAFF-R, TACI and APRIL in patients with autoimmune and/or oncologic diseases so that his role could be assessed. The study was made on Peripheral Blood (SP) of 11 healthy controls (CTL) and of 76 patients, 21 with autoimmune diseases (DAI), 47 with cancers (NEO) and 8 with both diseases (DAI + NEO). After the SP harvest we proceeded to the analysis of the different lymphocytic populations, with identification of the Treg cells and evaluation of the expression of BAFF, BAFF-R, TACI and APRIL by flow cytometry using fluorescent bind monoclonal antibodies. The results were statistically analyzed by Anova, T-student and multivariate analysis (p<0,05). Results: Upon the linfogram analysis, total lymphocytes from the DAI and NEO patients were decreased in comparison to the controls. It was observed a significant decline of the B lymphocytes on the DAI and DAI+NEO patients and an increase on T lymphocytes on DAI individuals when compared to healthy controls and NEO. The Treg cells of the controls represented only and small amount of cells, however the different groups of patients presented a significant increase of these cells percentage. Patients with cancer had an increase on BAFF expressing cells comparing to controls, on the other hand, DAI+NEO and DAI individuals showed diminished percentage of these cells. The surface receptor BAFF-R revealed biggest expression on DAI+NEO’s cells than in healthy controls’ cells. The TACI analysis showed that the group with oncologic disease had high expression that stood alone from the rest of the groups. Finally the APRIL ligand analysis revealed that DAI+NEO and NEO groups of patients had three times more expression of this molecule than the controls. Discussion: On the analyzed patients we verify an significant increase on Treg cells on DAI patients, individuals with oncologic disease and mainly on patients with both diseases, possibly a way of the oncologic pathologies create an immunossupressor environment and an attempt of the immune system of the DAI patients to suppress the abnormal inflammatory response. Considering the ligands BAFF, APRIL and the receptors BAFF-R and TACI, the group of oncologic patients revealed a substantial increase on their expression, disclosing that they can be acquired mechanisms of cellular survival in oncologic diseases. Conclusion: It was concluded that there are relevant disturbances on B and T lymphocytes, with change on their populations on both pathologies. Furthermore, our results suggest that the T regulatory cells and ligands and receptors related to NF-κB, BAFF, APRIL and BAFF-R and TACI, respectively, can participate on the fisiopathology of autoimmune and oncologic diseases, and eventually can compose new therapeutic targets to the treatment of these illnesse

Nontravel-related invasive Entamoeba histolytica infection with probable heterosexual transmission

Introduction: In industrialized countries, amebiasis usually occurs in migrants and travelers returning from areas where the disease is endemic, primarily by ingestion of contaminated food or water. Person-to-person transmission can occur, mainly by fecal-oral contact, but sexual transmission has also been described [1,3–5]. Presentation of case: We report a man with Entamoeba histolytica colitis and a large liver abscess (16.5 × 14 cm) in Portugal, who had no relevant travel history and whose only risk factor was his heterosexual partner. The abscess required drainage of 1950 mL of “chocolate-milk” purulent fluid, with rapid symptomatic improvement. The diagnosis was established by real-time reverse transcription PCR for Entamoeba histolytica in the liver aspirate, with positive IgG antibodies. He received a total of 16 days of ceftriaxone and metronidazole followed by 7 days of paromomycin. Conclusion: As enteric infections may be sexually transmitted, in industrialized countries, even in the absence of travel, sexual history should not be neglected. Keywords: Amebiasis, Entamoeba histolytica, Sexually Transmitted Diseases, Liver absces

Non-Ideality in Thymol + Menthol Type V Deep Eutectic Solvents

The nonionic Type V deep eutectic solvent (DESs) thymol + menthol is experimentally and computationally studied aiming to clarify the relation between its liquid phase structure and its thermodynamic nonideality. 1H NMR, Raman, and X-ray scattering analysis of the thymol + menthol system, supported by molecular dynamics simulations, show complex intermolecular interactions dominated by sterically hindered H-bonded clusters. For temperatures greater than or equal to room temperature, a quasi-linear evolution of the eutectic system properties between the pure compounds is observed, suggesting the absence of a magic stoichiometric composition in the eutectic solvent. However, temperature dependent Raman spectroscopy indicates a notable increase in thymol–menthol H-bonding as temperatures approach the eutectic point. This study shows that nonionic Type V DESs present an important temperature-dependent nonideality originating from the change in the intermolecular H-bonding with temperature. These findings have significant implications for the design and growing application of Type V DESs