The swiss transplant cohort study: lessons from the first 6 years.

Prospective cohort studies significantly contribute to answering specific research questions in a defined population. Since 2008, the Swiss Transplant Cohort Study (STCS) systematically enrolled >95 % of all transplant recipients in Switzerland, collecting predefined data at determined time points. Designed as an open cohort, the STCS has included >3900 patients to date, with a median follow-up of 2.96 years (IQR 1.44-4.73). This review highlights some relevant findings in the field of transplant-associated infections gained by the STCS so far. Three key general aspects have crystallized: (i) Well-run cohort studies are a powerful tool to conduct genetic studies, which are crucially dependent on a meticulously described phenotype. (ii) Long-term real-life observations are adding a distinct layer of information that cannot be obtained during randomized studies. (iii) The systemic collection of data, close interdisciplinary collaboration, and continuous analysis of some key outcome data such as infectious diseases endpoints can improve patient care.

The Effect of Everolimus Initiation and Calcineurin Inhibitor Elimination on Cardiac Allograft Vasculopathy in De Novo Recipients: One-Year Results of a Scandinavian Randomized Trial.

Early initiation of everolimus with calcineurin inhibitor therapy has been shown to reduce the progression of cardiac allograft vasculopathy (CAV) in de novo heart transplant recipients. The effect of de novo everolimus therapy and early total elimination of calcineurin inhibitor therapy has, however, not been investigated and is relevant given the morbidity and lack of efficacy of current protocols in preventing CAV. This 12-month multicenter Scandinavian trial randomized 115 de novo heart transplant recipients to everolimus with complete calcineurin inhibitor elimination 7-11 weeks after HTx or standard cyclosporine immunosuppression. Ninety-five (83%) patients had matched intravascular ultrasound examinations at baseline and 12 months. Mean (± SD) recipient age was 49.9 ± 13.1 years. The everolimus group (n = 47) demonstrated significantly reduced CAV progression as compared to the calcineurin inhibitor group (n = 48) (ΔMaximal Intimal Thickness 0.03 ± 0.06 and 0.08 ± 0.12 mm, ΔPercent Atheroma Volume 1.3 ± 2.3 and 4.2 ± 5.0%, ΔTotal Atheroma Volume 1.1 ± 19.2 mm(3) and 13.8 ± 28.0 mm(3) [all p-values ≤ 0.01]). Everolimus patients also had a significantly greater decline in levels of soluble tumor necrosis factor receptor-1 as compared to the calcineurin inhibitor group (p = 0.02). These preliminary results suggest that an everolimus-based CNI-free can potentially be considered in suitable de novo HTx recipients.

Immunologic basis of transplant-associated arteriosclerosis.

Although immunosuppressive therapy minimizes the risk of graft failure due to acute rejection, transplant-associated arteriosclerosis of the coronary arteries remains a significant obstacle to the long-term survival of heart transplant recipients. The participation of specific inflammatory cell types in the genesis of this lesion was examined in a mouse model in which carotid arteries were transplanted across multiple histocompatibility barriers into seven mutant strains with immunologic defects. An acquired immune response--with the participation of CD4+ (helper) T cells, humoral antibody, and macrophages--was essential to the development of the concentric neointimal proliferation and luminal narrowing characteristic of transplant arteriosclerosis. CD8+ (cytotoxic) T cells and natural killer cells were not involved in the process. Arteries allografted into mice deficient in both T-cell receptors and humoral antibody showed almost no neointimal proliferation, whereas those grafted into mice deficient only in helper T cells, humoral antibody, or macrophages developed small neointimas. These small neointimas and the large neointimas of arteries grafted into control animals contained a similar number of inflammatory cells; however, smooth muscle cell number and collagen deposition were diminished in the small neointimas. Also, the degree of inflammatory reaction in the adventitia did not correlate with the size of the neointima. Thus, the reduction in neointimal size in arteries allografted into mice deficient in helper T cells, humoral antibody, or macrophages may be accounted for by a decrease in smooth muscle cell migration or proliferation.

Area-under-the-curve monitoring of prednisolone for dose optimization in a stable renal transplant population

Background: Renal transplant recipients were noted to appear cushingoid while on low doses of steroid as part of a triple therapy immunosuppression of cyclosporin A (CsA), prednisolone, and azathioprine. Methods: The study group comprised adult renal transplant recipients with stable graft function who had received their renal allograft a minimum of 1 year previously (43 studies undertaken in 22 men and 20 women) with median daily prednisone dose of 7 mg (range 3-10). The control group was healthy nontransplant subjects [median dose 10 mg (10-30)]. Prednisolone bioavailability was measured using a limited 6-hour area under the curve (AUC), with prednisolone measured using specific HPLC assay. Results: The median prednisolone AUC/mg dose for all transplant recipients was significantly greater than the control group by approximately 50% (316 nmol(.)h/L/mg prednisolone versus 218). AUC was significantly higher in female recipients (median 415 versus 297 for men) and in recipients receiving cyclospotin (348 versus 285). The highest AUC was in women on estrogen supplements who were receiving cyclosporin (median 595). A significantly higher proportion of patients on triple therapy had steroid side effects compared with those on steroid and azathioprine (17/27 versus 4/15), more women than men had side effects (14/16 versus 7/22), and the AUC/mg prednisone was greater in those with side effects than without (median 377 versus 288 nmol-h/L/mg). Discussion: The results are consistent with the hypothesis that CsA increases the bioavailability of prednisolone, most likely through inhibition of beta-glycoprotein. The increased exposure to steroid increased the side-effect profile of steroids in the majority of patients. Because the major contributor to AUC is the maximum postdose concentration, it may be possible to use single-point monitoring (2 hours postdose) for routine clinical studies.

CEDIA (R) sirolimus assay compared with HPLC-MS/MS and HPLC-UV in transplant recipient specimens

The role of the therapeutic drug monitoring laboratory in support of immunosuppressant drug therapy is well established, and the introduction of sirolimus (SRL) is a new direction in this field. The lack of an immunoassay for several years has restricted the availability of SRL assay services. The recent availability of a CEDIA (R) SRL assay has the potential to improve this situation. The present communication has compared the CEDIA (R) SRL method with 2 established chromatographic methods, HPLC-UV and HPLC-MS/MS. The CEDIA (R) method, run on a Hitachi 917 analyzer, showed acceptable validation criteria with within-assay precision of 9.1% and 3.3%, and bias of 17.1% and 5.8%, at SRL concentrations of 5.0 mu g/L and 20 mu g/L, respectively. The corresponding between-run precision values were 11.5% and 3.3% and bias of 7.1% and 2.9% at 5.0 mu g/L and 20 mu g/L, respectively, The lower limit of quantification was found to be 3.0 mu g/L. A series of 96 EDTA whole-blood samples predominantly from renal transplant recipients were assayed by the 3 methods for comparison. It was found that the CEDIA (R) method showed a Deming regression line of CEDIA = 1.20 X HPLC-MS/MS - 0.07 (r = 0.934, SEE = 1.47), with a mean bias of 20.4%. Serial blood samples from 8 patients included in this evaluation showed that the CEDIA (R) method reflected the clinical fluctuations in the chromatographic methods, albeit with the variable bias noted. The CEDIA (R) method on the H917 analyzer is therefore a useful adjunct to SRL dosage individualization in renal transplant recipients.

Influence of ABCB1 polymorphisms and haplotypes on tacrolimus nephrotoxicity and dosage requirements in children with liver transplant

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT • Currently tacrolimus is the mainstay of immunosuppression for most children undergoing liver transplantation (LT). • The clinical use of this agent, however, is complicated by its various adverse effects (mainly nephrotoxicity), its narrow therapeutic-index and considerable pharmacokinetic variability. • The low and variable oral bioavailability of tacrolimus is thought to result from the action of the multidrug efflux-pump P-glycoprotein, encoded by the ABCB1 gene. WHAT THIS STUDY ADDS • A significant association between ABCB1 genetic polymorphisms and tacrolimus-associated nephrotoxicity in paediatric patients following LT is reported for the first time. Genotyping such polymorphisms may have the potential to individualize better initial tacrolimus therapy and enhance drug safety. • The long-term effect of ABCB1 polymorphisms on tacrolimus trough concentrations were investigated up to 5 years post-transplantation. A significant effect of intestinal P-glycoprotein genotypes on tacrolimus pharmacokinetics was found at 3 and 4 years post-transplantation suggesting that the effect is maintained long term. AIMS - The aim of this study was to investigate the influence of genetic polymorphisms in ABCB1 on the incidence of nephrotoxicity and tacrolimus dosage-requirements in paediatric patients following liver transplantation. METHODS - Fifty-one paediatric liver transplant recipients receiving tacrolimus were genotyped for ABCB1 C1236>T, G2677>T and C3435>T polymorphisms. Dose-adjusted tacrolimus trough concentrations and estimated glomerular filtration rates (EGFR) indicative of renal toxicity were determined and correlated with the corresponding genotypes. RESULTS - The present study revealed a higher incidence of the ABCB1 variant-alleles examined among patients with renal dysfunction (≥30% reduction in EGFR) at 6 months post-transplantation (1236T allele: 63.3% vs 37.5% in controls, P= 0.019; 2677T allele: 63.3% vs. 35.9%, p = 0.012; 3435T allele: 60% vs. 39.1%, P= 0.057). Carriers of the G2677->T variant allele also had a significant reduction (%) in EGFR at 12 months post-transplant (mean difference = 22.6%; P= 0.031). Haplotype analysis showed a significant association between T-T-T haplotypes and an increased incidence of nephrotoxicity at 6 months post-transplantation (haplotype-frequency = 52.9% in nephrotoxic patients vs 29.4% in controls; P= 0.029). Furthermore, G2677->T and C3435->T polymorphisms and T-T-T haplotypes were significantly correlated with higher tacrolimus dose-adjusted pre-dose concentrations at various time points examined long after drug initiation. CONCLUSIONS - These findings suggest that ABCB1 polymorphisms in the native intestine significantly influence tacrolimus dosage-requirement in the stable phase after transplantation. In addition, ABCB1 polymorphisms in paediatric liver transplant recipients may predispose them to nephrotoxicity over the first year post-transplantation. Genotyping future transplant recipients for ABCB1 polymorphisms, therefore, could have the potential to individualize better tacrolimus immunosuppressive therapy and enhance drug safety.

PULMONARY REHABILITATION PROGRAMS IN LUNG TRANSPLANT: A LITERATURE REVIEW

Objective: To analyze, using a literature review, Pulmonary Rehabilitation (RP) Programs in lung transplant. Methods: A literature review in July 2014 in Ebsco Host, Periódicos Capes, BVS and Science Direct data bases using descriptors in English (“lung transplantation”, “lung transplant” AND/OR “rehabilitation”) and Portuguese (“reabilitação” AND/OR “transplante pulmonar”). The eligibility criterions were interventional studies of PR before and/or after lung transplant; participants who were candidates to lung transplant or lung transplant recipients; studies that applied any kind of PR program (hospital-based, homebased or outpatient) and articles published in English, Spanish or Portuguese. Literature reviews, guidelines and case reports were excluded. The search process yielded 46 articles of which two were duplicated. After title and abstract screening 13 articles remained for full text reading. Six studies met the inclusion eligibility and were included in the review. Results: The studies involved patients with Chronic Obstructive Pulmonary Disease, Cystic Fibrosis, Pulmonary Hypertension, Interstitial Lung Disease and Pulmonary Fibrosis. Pulmonary function, exercise capacity, quality of life (QoL) and quadriceps force were evaluated. Most interventions were outpatient programs with three months duration, three times a week and session with at least one hour. Protocols included physical training, educational approach and just one included nutritional, psychiatric and social assistant follow-up. The studies presented significant change in the six-minute walking distance, QoL and quadriceps force after PR programs. Conclusion: This review showed the benefits of the PR in the QoL and exercise capacity contributing to the Health Promotion of the patients.

Prevention of transnational transplant-related crimes—What more can be done?

Many nations are able to prosecute transplant-related crimes committed in their territory, but transplant recipients, organ sellers and brokers, and transplant professionals may escape prosecution by engaging in these practices in foreign locations where they judge the risk of criminal investigation and prosecution to be remote.Methods. The Declaration of Istanbul Custodian Group convened an international working group to evaluate the possible role of extraterritorial jurisdiction in strengthening the enforcement of existing laws governing transplant-related crimes across national boundaries. Potential practical and ethical concerns about the use of extraterritorial jurisdiction were examined, and possible responses were explored. Results. Extraterritorial jurisdiction is a legitimate tool to combat transplant-related crimes. Further, development of a global registry of transnational transplant activities in conjunction with a standardized international referral system for legitimate travel for transplantation is proposed as a mechanism to support enforcement of national and international legal tools. Conclusions. States are encouraged to include provisions on extraterritorial jurisdiction in their laws on transplant-related crimes and to collaborate with professionals and international authorities in the development of a global registry of transnational transplant activities. These actions would assist in the identification and evaluation of illicit activities and provide information that would help in developing strategies to deter and prevent them.

Determining the psychosocial predictors of living, living-related, and posthumous organ donation

The worldwide organ shortage occurs despite people’s positive organ donation attitudes. The discrepancy between attitudes and behaviour is evident in Australia particularly, with widespread public support for organ donation but low donation and communication rates. This problem is compounded further by the paucity of theoretically based research to improve our understanding of people’s organ donation decisions. This program of research contributes to our knowledge of individual decision making processes for three aspects of organ donation: (1) posthumous (upon death) donation, (2) living donation (to a known and unknown recipient), and (3) providing consent for donation by communicating donation wishes on an organ donor consent register (registering) and discussing the donation decision with significant others (discussing). The research program used extended versions of the Theory of Planned Behaviour (TPB) and the Prototype/Willingness Model (PWM), incorporating additional influences (moral norm, self-identity, organ recipient prototypes), to explicate the relationship between people’s positive attitudes and low rates of organ donation behaviours. Adopting the TPB and PWM (and their extensions) as a theoretical basis overcomes several key limitations of the extant organ donation literature including the often atheoretical nature of organ donation research, thefocus on individual difference factors to construct organ donor profiles and the omission of important psychosocial influences (e.g., control perceptions, moral values) that may impact on people’s decision-making in this context. In addition, the use of the TPB and PWM adds further to our understanding of the decision making process for communicating organ donation wishes. Specifically, the extent to which people’s registering and discussing decisions may be explained by a reasoned and/or a reactive decision making pathway is examined (Stage 3) with the novel application of the TPB augmented with the social reaction pathway in the PWM. This program of research was conducted in three discrete stages: a qualitative stage (Stage 1), a quantitative stage with extended models (Stage 2), and a quantitative stage with augmented models (Stage 3). The findings of the research program are reported in nine papers which are presented according to the three aspects of organ donation examined (posthumous donation, living donation, and providing consent for donation by registering or discussing the donation preference). Stage One of the research program comprised qualitative focus groups/interviews with university students and community members (N = 54) (Papers 1 and 2). Drawing broadly on the TPB framework (Paper 1), content analysed responses revealed people’s commonly held beliefs about the advantages and disadvantages (e.g., prolonging/saving life), important people or groups (e.g., family), and barriers and motivators (e.g., a family’s objection to donation), related to living and posthumous organ donation. Guided by a PWM perspective, Paper Two identified people’s commonly held perceptions of organ donors (e.g., altruistic and giving), non-donors (e.g., self-absorbed and unaware), and transplant recipients (e.g., unfortunate, and in some cases responsible/blameworthy for their predicament). Stage Two encompassed quantitative examinations of people’s decision makingfor living (Papers 3 and 4) and posthumous (Paper 5) organ donation, and for registering and discussing donation wishes (Papers 6 to 8) to test extensions to both the TPB and PWM. Comparisons of health students’ (N = 487) motivations and willingness for living related and anonymous donation (Paper 3) revealed that a person’s donor identity, attitude, past blood donation, and knowing a posthumous donor were four common determinants of willingness, with the results highlighting students’ identification as a living donor as an important motive. An extended PWM is presented in Papers Four and Five. University students’ (N = 284) willingness for living related and anonymous donation was tested in Paper Four with attitude, subjective norm, donor prototype similarity, and moral norm (but not donor prototype favourability) predicting students’ willingness to donate organs in both living situations. Students’ and community members’ (N = 471) posthumous organ donation willingness was assessed in Paper Five with attitude, subjective norm, past behaviour, moral norm, self-identity, and prior blood donation all significantly directly predicting posthumous donation willingness, with only an indirect role for organ donor prototype evaluations. The results of two studies examining people’s decisions to register and/or discuss their organ donation wishes are reported in Paper Six. People’s (N = 24) commonly held beliefs about communicating their organ donation wishes were explored initially in a TPB based qualitative elicitation study. The TPB belief determinants of intentions to register and discuss the donation preference were then assessed for people who had not previously communicated their donation wishes (N = 123). Behavioural and normative beliefs were important determinants of registering and discussing intentions; however, control beliefs influenced people’s registering intentions only. Paper Seven represented the first empirical test of the role of organ transplant recipient prototypes (i.e., perceptions of organ transplant recipients) in people’s (N = 465) decisions to register consent for organ donation. Two factors, Substance Use and Responsibility, were identified and Responsibility predicted people’s organ donor registration status. Results demonstrated that unregistered respondents were the most likely to evaluate transplant recipients negatively. Paper Eight established the role of organ donor prototype evaluations, within an extended TPB model, in predicting students’ and community members’ registering (n = 359) and discussing (n = 282) decisions. Results supported the utility of an extended TPB and suggested a role for donor prototype evaluations in predicting people’s discussing intentions only. Strong intentions to discuss donation wishes increased the likelihood that respondents reported discussing their decision 1-month later. Stage Three of the research program comprised an examination of augmented models (Paper 9). A test of the TPB augmented with elements from the social reaction pathway in the PWM, and extensions to these models was conducted to explore whether people’s registering (N = 339) and discussing (N = 315) decisions are explained via a reasoned (intention) and/or social reaction (willingness) pathway. Results suggested that people’s decisions to communicate their organ donation wishes may be better explained via the reasoned pathway, particularly for registering consent; however, discussing also involves reactive elements. Overall, the current research program represents an important step toward clarifying the relationship between people’s positive organ donation attitudes but low rates of organ donation and communication behaviours. Support has been demonstrated for the use of extensions to two complementary theories, the TPB and PWM, which can inform future research aiming to explicate further the organ donation attitude-behaviour relationship. The focus on a range of organ donation behaviours enables the identification of key targets for future interventions encouraging people’s posthumous and living donation decisions, and communication of their organ donation preference.

Myeloma-induced alloreactive T cells arising in myeloma-infiltrated bones include double-positive CD8+CD4+ T cells : evidence from myeloma-bearing mouse model

The graft-versus-myeloma (GVM) effect represents a powerful form of immune attack exerted by alloreactive T cells against multiple myeloma cells, which leads to clinical responses in multiple myeloma transplant recipients. Whether myeloma cells are themselves able to induce alloreactive T cells capable of the GVM effect is not defined. Using adoptive transfer of T naive cells into myeloma-bearing mice (established by transplantation of human RPMI8226-TGL myeloma cells into CD122(+) cell-depleted NOD/SCID hosts), we found that myeloma cells induced alloreactive T cells that suppressed myeloma growth and prolonged survival of T cell recipients. Myeloma-induced alloreactive T cells arising in the myeloma-infiltrated bones exerted cytotoxic activity against resident myeloma cells, but limited activity against control myeloma cells obtained from myeloma-bearing mice that did not receive T naive cells. These myeloma-induced alloreactive T cells were derived through multiple CD8(+) T cell divisions and enriched in double-positive (DP) T cells coexpressing the CD8alphaalpha and CD4 coreceptors. MHC class I expression on myeloma cells and contact with T cells were required for CD8(+) T cell divisions and DP-T cell development. DP-T cells present in myeloma-infiltrated bones contained a higher proportion of cells expressing cytotoxic mediators IFN-gamma and/or perforin compared with single-positive CD8(+) T cells, acquired the capacity to degranulate as measured by CD107 expression, and contributed to an elevated perforin level seen in the myeloma-infiltrated bones. These observations suggest that myeloma-induced alloreactive T cells arising in myeloma-infiltrated bones are enriched with DP-T cells equipped with cytotoxic effector functions that are likely to be involved in the GVM effect.

Cytomegalovirus in the Process of Chronic Allograft Nephropathy

Cytomegalovirus (CMV) is a major cause of morbidity, costs and even mortality in organ transplant recipients. CMV may also enhance the development of chronic allograft nephropathy (CAN), which is the most important cause of graft loss after kidney transplantation. The evidence for the role of CMV in chronic allograft nephropathy is somewhat limited, and controversial results have also been reported. The aim of this study was to investigate the role of CMV in the pathogenesis of CAN. Material for the purpose of this study was available from altogether 70 kidney transplant recipients who received a kidney transplant between the years 1992-2000. CMV infection was diagnosed with pp65 antigenemia test or by viral culture from blood, urine, or both. CMV proteins were demonstrated in the kidney allograft biopsies by immunohistochemisrty and CMV-DNA by in situ hybridization. Cytokines, adhesion molecules, and growth factors were demonstrated from allograft biopsies by immunohistochemistry, and from urinary samples by ELISA-methods. CMV proteins were detectable in the 6-month protocol biopsies from 18/41 recipients with evidence of CMV infection. In the histopathological analysis of the 6-month protocol biopsies, presence of CMV in the allograft together with a previous history of acute rejection episodes was associated with increased arteriosclerotic changes in small arterioles. In urinary samples collected during CMV infection, excretion of TGF-β was significantly increased. In recipients with increased urinary excretion of TGF-β, increased interstitial fibrosis was recorded in the 6- month protocol biopsies. In biopsies taken after an active CMV infection, CMV persisted in the kidney allograft in 17/48 recipients, as CMV DNA or antigens were detected in the biopsies more than 2 months after the last positive finding in blood or urine. This persistence was associated with increased expression of TGF-β, PDGF, and ICAM-1 and with increased vascular changes in the allografts. Graft survival and graft function one and two years after transplantation were reduced in recipients with persistent intragraft CMV. Persistent intragraft CMV infection was also a risk factor for reduced graft survival in Cox regression analysis, and an independent risk factor for poor graft function one and two years after transplantation in logistic regression analysis. In conclusion, these results show that persistent intragraft CMV infection is detrimental to kidney allografts, causing increased expression of growth factors and increased vascular changes, leading to reduced graft function and survival. Effective prevention, diagnosis and treatment of CMV infections may a major factor in improving the long term survival of kidney allograft.

Matrix metalloproteinases as biomarkers in premalignant and malignant tumors of the human skin

The incidence of non-melanoma skin cancer is increasing worldwide. Basal cell carcinoma followed by squamous cell carcinoma and malignant melanoma are the most frequent skin tumors. Immunosuppressed patients have an increased risk of neoplasia, of which non-melanoma skin cancer is the most common. Matrix metalloproteinases (MMPs) are proteolytic enzymes that collectively are capable of degrading virtually all components of the extracellular matrix. MMPs can also process substrates distinct from extracellular matrix proteins and influence cell proliferation, differentiation, angiogenesis, and apoptosis. MMP activity is regulated by their natural inhibitors, tissue inhibitors of metallopro-teinases (TIMPs). In this study, the expression patterns of MMPs, TIMPs, and certain cancer-related molecules were investigated in premalignant and malignant lesions of the human skin. As methods were used immunohistochemisty, in situ hybridization, and reverse transcriptase polymerase chain reaction (RT-PCR) from the cell cultures. Our aim was to evaluate the expression pattern of MMPs in extramammary Paget's disease in order to find markers for more advanced tumors, as well as to shed light on the origin of this rare neoplasm. Novel MMPs -21, -26, and -28 were studied in melanoma cell culture, in primary cutaneous melanomas, and their sentinel nodes. The MMP expression profile in keratoacanthomas and well-differentiated squamous cell carcinomas was analyzed to find markers to differentiate benign keratinocyte hyperproliferation from malignantly transformed cells. Squamous cell carcinomas of immunosuppressed organ transplant recipients were compared to squamous cell carcinomas of matched immunocompetent controls to investigate the factors explaining their more aggressive nature. We found that MMP-7 and -19 proteins are abundant in extramammary Paget's disease and that their presence may predict an underlying adenocarcinoma in these patients. In melanomas, MMP-21 was upregulated in early phases of melanoma progression, but disappeared from the more aggressive tumors with lymph node metastases. The presence of MMP-13 in primary melanomas and lymph node metastases may relate to more aggressive disease. In keratoacanthomas, the expression of MMP-7 and -9 is rare and therefore should raise a suspicion of well-differentiated squamous cell carcinomas. Furthermore, MMP-19 and p16 were observed in benign keratinocyte hyperproliferation of keratoacanthomas, whereas they were generally lost from malignant keratinocytes of SCCs. MMP-26 staining was significantly stronger in squamous cell carcinomas and Bowen s disease samples of organ transplant recipients and it may contribute to the more aggressive nature of squamous cell carcinomas in immunosuppressed patients. In addition, the staining for MMP-9 was significantly stronger in macrophages surrounding the tumors of the immunocompetent group and in neutrophils of those patients on cyclosporin medication. In conclusion, based on our studies, MMP-7 and -19 might serve as biomarkers for more aggressive extramammary Paget's disease and MMP-21 for malignant transformation of melanocytes. MMP -7, -9, and -26, however, could play an important role in the pathobiology of keratinocyte derived malignancies.

Estudo do metabolismo mitocondrial e da resposta anti-apoptótica de células endoteliais humanas durante a evolução da infecção por taquizoítos de Toxoplasma gondii

A toxoplasmose é uma zoonose amplamente distribuída que afeta mais de um terço da população mundial e de grande importância na saúde pública. A maioria das infecções em humanos por Toxoplasma gondii é assintomática. A toxoplasmose é amplamente investigada visto que se apresenta como uma doença grave em pessoas imunodeprimidas (portadores da síndrome da imunodeficiência adquirida (SIDA), não tratados, indivíduos transplantados, paciente em tratamento quimioterápico ou em uso de drogas supressoras e gestantes). A toxoplasmose congênita frequentemente pode levar ao aborto espontâneo ou até mesmo resultar na formação de crianças com algum grau de atraso no desenvolvimento mental e/ou físicos, deste modo, a transmissão congênita pode ser muito mais importante do que se pensava, pois os parasitos encontrados na circulação sanguinea são capazes de infectar as células endoteliais dos vasos e os tecidos circunjacentes, podendo resultar no encistamento do T. gondii. Atualmente a toxoplasmose vem sendo investigada devido a sua associação a inúmeras outras doenças, assim, estudos sobre a evolução da infecção por T. gondii em diferentes tipos de células hospedeiras se fazem necessários para uma abordagem terapêutica adequada. Ao invadir a célula hospedeira o parasito possui a capacidade de recrutar as mitocôndrias promovendo mudanças na organização mitocondrial ao longo da progressão da infecção, garantindo um ambiente favorável a sua multiplicação. Diante disso, investigamos se o parasito possui a capacidade de interferir no metabolismo mitocondrial e na resposta apoptótica da célula endotelial. O presente trabalho teve como objetivo analisar o metabolismo mitocondrial através da respirometria de alta-resolução e da resposta apoptótica através do western blotting das células endoteliais da veia umbilical humana (HUVEC) infectadas por 2, 6 e 20 horas por taquizoítos de T. gondii. A respirometria de alta-resolução revelou que o parasito interfere no metabolismo energético da célula hospedeira. A análise do conteúdo de proteínas da família Bcl-2 por western blotting revelou maior estímulo apoptótico no tempo inicial de infecção, quando comparado aos demais tempos. Os resultados dos conteúdos de caspase 3, proteína efetora da apoptose, não demonstrou diferença nos tempos iniciais de infecção Entretanto, em tempos mais tardios, o conteúdo de caspase 3 mostrou-se significativamente aumentado quando comparado às HUVEC não infectadas. A dinâmica de replicação do parasito foi observada através do monitoramento pelo sistema Time-Lapse Nikon BioStation IMQ em tempo real das células infectadas por T.gondii. Portanto, nossos resultados sugerem que o protozoário ao recrutar as mitocôndrias da célula hospedeira interfere no metabolismo mitocondrial e na modulação da apoptose para garantir um ambiente favorável a sua multiplicação.

Quorum Sensing na Prática Clínica

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas