Genetic polymorphisms of toll-like receptors in leprosy patients from southern Brazil.

Leprosy is a chronic disease and also a global health issue, with a high number of new cases per year. Toll-like receptors can respond to mycobacterial molecules in the early stage of infection. As important components of the innate immune response, alterations in genes coding for these receptors may contribute to susceptibility/protection against diseases. In this context, we used a case-control study model (183 leprosy cases vs. 185 controls) to investigate whether leprosy patients and the control group, in southern Brazil, have different frequencies in TLR1 (TLR1 G>T; rs5743618), TLR2 (TLR2 T>C, rs1816702 and rs4696483), and TLR4 (TLR4 A>G, rs1927911) polymorphisms. Analysis of the TLR1 1805G>T polymorphism presented the G/G genotype more frequently in the control group. TLR2 T>C rs1816702 and TLR2 T>C rs4696483, the T/T and C/T genotype, respectively, were more frequent in the control group than in leprosy patients, suggesting protection from leprosy when the T allele is present (rs4696483). Haplotype analyses between TLR1 (rs5743618) and TLR2 (rs1816702 and rs4696483) polymorphisms suggest risk for the presence of the TCC haplotype and protection in the presence of the TCT haplotype. This study suggests that polymorphisms in TLR1 and TLR2 are factors that may contribute to development/resistance of leprosy.

NKT-Like (CD3+CD56+) Cells in Chronic Myeloid Leukemia Patients Treated With Tyrosine Kinase Inhibitors.

Therapy with Tyrosine Kinase Inhibitors (TKI) aiming stable deep molecular response is the gold standard to treat Chronic Myeloid Leukemia (CML). NKT-like cells (CD3+CD56+) combine characteristics of T and NK cells. The physiopathological role of these cells remains unknown although the literature refers their association with inflammation, autoimmune diseases, and cancer. Since the information regarding the role of NKT-like cells in CML is rare, we aimed at the characterization of these cells in CML patients treated with TKIs. Peripheral blood NKT-like cells from 48 CML patients and 40 healthy donors were analyzed by multiparametric flow cytometry. Functional tests consisting of co-culture with leukemic target cells (K562 cell line) were used to measure degranulation and cytokine production. Our results revealed that NKT-like cells are decreased in treated CML patients, although they present increased expression of activation markers (CD69 and HLA-DR), increased degranulation (CD107a) and impaired IFN-γ production. Significantly alterations on the expression of tumor recognition (NCRs and NKp80), and immune regulation receptors (LAG-3, TIM-3, and CD137) by NKT-like cells were observed in CML patients. Second generation TKIs increased cell activation (CD69) and decreased expression of NKp44 and NKp80 by NKT-like cells from CML patients when compared to Imatinib. CML patients that achieved deep molecular response (MR4.5) presented downregulation of NKp44 and LAG-3. Further studies are needed to clarify the role of these cells as biomarkers of therapy response and also to evaluate their value for discrimination of better candidates for sustained treatment-free remission after TKI discontinuation.

Development of a Healthy Lifestyle Assessment Toolkit for the General Public.

The prevalence of age-related non-communicable chronic diseases has increased worldwide, being the leading causes of morbidity and death in many world regions, including in Europe. Innovative models and strategies focused on preventive care, including early identification of risk factors underlying disease onset and progression, and proper modification of lifestyle habits and behaviors, might contribute to promote quality of life, healthy living and active aging. Healthy Lifestyle Innovative Quarters for Cities and Citizens (HeaLIQs4cities) is an EIT Health-funded project aiming to engage, empower and educate citizens toward healthy lifestyles. One of the major objectives of this project was to develop a toolkit for a rapid and informal assessment of healthy lifestyles, to be used at different levels of care pathways, including in informal public environments. In this paper, we describe the methodology underlying the development of the toolkit, which resulted from the collaboration of an interdisciplinary focus group of academic experts, from medicine, sport sciences, psychology, health economics, and innovative technologies applied to health. The following eight components were included in the toolkit: (1) anthropometric assessment and cardiometabolic parameters; (2) physical activity and exercise; (3) well-being, social cohesion, and functional independence; (4) nutrition; (5) mental health; (6) smoking, drinking, and use of illicit substances; (7) sleep habits and quality; and (8) health and disease. A traffic light rating system indicating the risk score was used (low: green; moderate: yellow; and relevant: orange) for each of the 8 components, together with recommendations for the toolkit users. After completing the reduced version of the toolkit, individuals showing moderate or relevant risk in one or more of the 8 dimensions, were invited to participate in a more detailed assessment (toolkit long version), based on deeper and scientifically validated tools. The toolkit was incorporated in eVida, a web-based platform that focuses on delivering services to personalized health and well-being. The validation of the current toolkit has been applied in wide-ranging public events in three different European Regions. Large scale deployment of the toolkit is expected to profit from the Reference Site Collaborative Network of the European Innovation Partnership on Active and Healthy Aging (EIP on AHA).

Effect of Age on NK Cell Compartment in Chronic Myeloid Leukemia Patients Treated With Tyrosine Kinase Inhibitors.

Natural killer (NK) cells are a very important component of the innate immune response involved in the lysis of virus infected and tumor cells. Aging has a profound impact in the frequency, phenotype and function of NK cells. Chronic Myeloid Leukemia (CML) is caused by the BCR-ABL gene formation encoding aberrant oncoprotein tyrosine kinase. Treatment with tyrosine kinase inhibitors (TKIs) induces durable deep molecular response. The response to treatment and life expectancy is lower in older patients with chronic phase of CML than in younger patients. In this work we analyse NK cells from TKI-treated CML patients and healthy controls stratified according to age. We have analyzed the expression of NK receptors, activation markers, NK cell differentiation in CD56bright and CD56dim NK cell subsets and the expression of CD107a and IFN-γ in NK cells stimulated with K562. Whereas significant differences on the phenotype and function of NK cells were found between middle-aged (35-65 years old) and elderly (older than 65) healthy individuals, NK cells from TKI-treated CML patients do not show significant differences related with age in most parameters studied, indicating that age is not a limitation of the NK cell recovery after treatment with TKI. Our results also revealed differences in the expression of NK receptors, activation markers and functional assays in NK cells from TKI-treated CML patients compared with age-matched healthy controls. These results highlight the relevance of NK cells in TKI-treated patients and the need of an extensive analysis of the effect of aging on NK cell phenotype and function in these patients in order to define new NK-cell based strategies directed to control CML progression and achieve long-term disease remission after TKI cessation.

The effects of physical exercise on nonmotor symptoms and on neuroimmune RAGE network in experimental parkinsonism.

Parkinson's disease (PD) prodromal stages comprise neuropsychiatric perturbations that critically compromise a patient's quality of life. These nonmotor symptoms (NMS) are associated with exacerbated innate immunity, a hallmark of overt PD. Physical exercise (PE) has the potential to improve neuropsychiatric deficits and to modulate immune network including receptor for advanced glycation end products (RAGE) and Toll-like receptors (TLRs) in distinct pathological settings. Accordingly, the present study aimed to test the hypothesis that PE 1) alleviates PD NMS and 2) modulates neuroimmune RAGE network in experimental PD. Adult Wistar rats subjected to long-term mild treadmill were administered intranasally with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and probed for PD NMS before the onset of motor abnormalities. Twelve days after MPTP, neuroimmune RAGE network transcriptomics (real-time quantitative PCR) was analyzed in frontal cortex, hippocampus, and striatum. Untrained MPTP animals displayed habit-learning and motivational deficits without gross motor impairments (cued version of water-maze, splash, and open-field tests, respectively). A suppression of RAGE and neuroimmune-related genes was observed in frontal cortex on chemical and physical stressors (untrained MPTP: RAGE, TLR5 and -7, and p22 NADPH oxidase; saline-trained animals: RAGE, TLR1 and -5 to -11, TNF-α, IL-1ß, and p22 NADPH oxidase), suggesting the recruitment of compensatory mechanisms to restrain innate inflammation. Notably, trained MPTP animals displayed normal cognitive/motivational performances. Additionally, these animals showed normal RAGE expression and neuroprotective PD-related DJ-1 gene upregulation in frontal cortex when compared with untrained MPTP animals. These findings corroborate PE efficacy in improving PD NMS and newly identify RAGE network as a neural substrate for exercise intervention. Additional research is warranted to unveil functional consequences of PE-induced modulation of RAGE/DJ-1 transcriptomics in PD premotor stages.NEW & NOTEWORTHY This study newly shows that physical exercise (PE) corrects nonmotor symptoms of the intranasal 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of experimental parkinsonism. Additionally, we show that suppression of neuroimmune receptor for advanced glycation end products (RAGE) network occurs in frontal cortex on chemical (MPTP) and physical (PE) interventions. Finally, PE normalizes frontal cortical RAGE transcriptomics and upregulates the neuroprotective DJ-1 gene in the intranasal MPTP model of experimental parkinsonism.

Regulation of striatal astrocytic receptor for advanced glycation end-products variants in an early stage of experimental Parkinson's disease.

Convincing evidence indicates that advanced glycation end-products and danger-associated protein S100B play a role in Parkinson's disease (PD). These agents operate through the receptor for advanced glycation end-products (RAGE), which displays distinct isoforms playing protective/deleterious effects. However, the nature of RAGE variants has been overlooked in PD studies. Hence, we attempted to characterize RAGE regulation in early stages of PD striatal pathology. A neurotoxin-based rodent model of PD was used in this study, through administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to C57BL/6 mice. Animals were killed 6 h post-MPTP to assess S100B/RAGE contents (RT-qPCR, ELISA) and RAGE isoform density (WB) and cellular distribution (immunohistochemistry). Dopaminergic and gliotic status were also mapped (HPLC-ED, WB, immunohistochemistry). At this preliminary stage of MPTP-induced PD in mice, RAGE inhibitory isoforms were increased whereas full-length RAGE was not affected. This putative cytoprotective RAGE phenotype paired an inflammatory and pro-oxidant setting fueling DAergic denervation. Increased RAGE inhibitory variants occur in astrocytes showing higher S100B density but no overt signs of hypertrophy or NF-κB activation, a canonical effector of RAGE. These findings expand our understanding of the toxic effect of MPTP on striatum and offer first in vivo evidence of RAGE being a responder in early stages of astrogliosis dynamics, supporting a protective rather tissue-destructive phenotype of RAGE in the initial phase of PD degeneration. These data lay the groundwork for future studies on the relevance of astrocytic RAGE in DAergic neuroprotection strategies. We report increased antagonistic RAGE variants paralleling S100B up-regulation in early stages of MPTP-induced astrogliosis dynamics . We propose that selective RAGE regulation reflects a self-protective mechanism to maintain low levels of RAGE ligands , preventing long-term inflammation and oxidative stress arising from sustained ligands/flRAGE activation . Understanding loss of RAGE protective response to stress may provide new therapeutic options to halt or slow down dopaminergic axonopathy and, ultimately, neuronal death .

EGFR/erB-1, HER2/erB-2, CK7, LP34, Ki67 and P53 expression in preneoplastic lesions of bronchial epithelium: an immunohistochemical and genetic study.

A prognostic interpretation of preneoplastic lesions would have impact in bronchial carcinoma early diagnosis and through the study of Erb-B family receptors as they have an important role in lung carcinogenesis. The existence of drugs as tyrosine kinase inhibitors stressed the importance of studying gene alterations for selected chemoprevention schemes and characterization of carcinogenesis. Bronchial preneoplastic lesions were characterized by immunohistochemistry using the antibodies LP34 (high weigh molecular cytokeratin), CK7, chromogranin A, Ki67, p53, C-erbB-2 and EGFR. HER2 and EGFR gene copy number was also evaluated by fluorescent in situ hybridization in those lesions. The expected results defined the origin cell for basal cell hyperplasia and squamous metaplasia as adaptative lesions and dysplasia. By known experiences and published data, beyond the stem cell, the spectral evolution of bronchial preneoplastic lesions was demonstrated by characterizing basal cells (LP34) and their neoplastic potentiality. Dysplasias showed a higher expression of EGFR, Ki67 and p53 with a stepwise increase with the gravity of the respective grading. C-erbB-2 immunohistochemical overexpression was a rare event in preneoplastic lesions. Polysomy was the main mechanism for EGFR and HER2/neu higher gene copy number and together with increased proliferation index (Ki67) will account to preview bronchial carcinogenesis.

Polysomy and amplification of chromosome 7 defined for EGFR gene in squamous cell carcinoma of the lung together with exons 19 and 21 wild type.

PURPOSE: The epidermal growth factor receptor (EGFR) is overexpressed in the majority of nonsmall- cell lung cancers (NSCLC) and is a major target specific EGFR tyrosine kinase inhibitors (TKIs) developed and used for the treatment of advanced NSCLC. A number of biological factors are also associated with EGFR-TKIs responsiveness. This study was focused on EGFR somatic mutations and amplifications in squamous cell lung cancer. MATERIAL AND METHODS: Representative sections of squamous cell carcinoma were selected from 54 surgical specimens from formalin-fixed paraffin-embedded tissues and submitted to TMA construction. Determination of EGFR protein expression was done by immunohistochemistry( IHC) (Zymed, Laboratories). Fluorescence in situ hybridization (FISH) was performed with LSI EGFR/CEP 7 (Vysis; Abbott Molecular, USA). Genomic DNA was extracted from 48 cases and exon 19 was amplified by polymerase chain reaction (PCR) for search deletions and point mutations for exon 21. All cases expressed high weigh cytokeratin and were observed negativity for CK7, CD56 and chromogranin. RESULTS: EGFR protein overexpression was identified in 49 cases, by the application of Hirsh's scoring system. The chromosome 7 and EGFR gene were analyzed by FISH and scored according to Cappuzzo's method that showed high polysomy in 31 cases and amplification in 7 cases. Deletion in exon 19 of EGFR was detected in 3 cases of 48 samples; the exon 21 of EGFR was expressed in its wild type by RFLP in all cases. CONCLUSIONS: Detection of common EGFR deletion and mutation showed to be a rare event in Squamous cell carcinoma of the lung. While EGFR mutation is the most effective molecular predictor or sensitivity in patients with advanced NSCLC submitted to EGFR-TKIs treatment, amplification and polysomy is the most effective molecular predictor for EGFR-TKIs responsiveness in squamous cell carcinoma, when validated isolated from the group of NSCLC.

Nitric oxide stimulates the proliferation of neural stem cells bypassing the epidermal growth factor receptor.

Nitric oxide (NO) was described to inhibit the proliferation of neural stem cells. Some evidence suggests that NO, under certain conditions, can also promote cell proliferation, although the mechanisms responsible for a potential proliferative effect of NO in neural stem cells have remained unaddressed. In this work, we investigated and characterized the proliferative effect of NO in cell cultures obtained from the mouse subventricular zone. We found that the NO donor NOC-18 (10 microM) increased cell proliferation, whereas higher concentrations (100 microM) inhibited cell proliferation. Increased cell proliferation was detected rapidly following exposure to NO and was prevented by blocking the mitogen-activated kinase (MAPK) pathway, independently of the epidermal growth factor (EGF) receptor. Downstream of the EGF receptor, NO activated p21Ras and the MAPK pathway, resulting in a decrease in the nuclear presence of the cyclin-dependent kinase inhibitor 1, p27(KIP1), allowing for cell cycle progression. Furthermore, in a mouse model that shows increased proliferation of neural stem cells in the hippocampus following seizure injury, we observed that the absence of inducible nitric oxide synthase (iNOS(-/-) mice) prevented the increase in cell proliferation observed following seizures in wild-type mice, showing that NO from iNOS origin is important for increased cell proliferation following a brain insult. Overall, we show that NO is able to stimulate the proliferation of neural stem cells bypassing the EGF receptor and promoting cell division. Moreover, under pathophysiological conditions in vivo, NO from iNOS origin also promotes proliferation in the hippocampus.

[The IASLC lung cancer staging project. Comparing the current 6(th) TNM edition with the proposed 7(th) edition]. / Projecto de estadiamento do cancro do pulmão pela IASLC: Estudo comparativo entre a 6.a edição TNM em vigor e a 7.a edição proposta.

The future 7th edition of TNM classification for lung cancer will be published in 2009 and comprises the IASLC recommendations for TNM parameters. The general staging of lung cancer includes the new parameters: reclassification of tumours larger than 7 cm from T2 to T3; extra tumoral nodules will change their category to T3, T4 and M1 when in the same, ipsilateral or contralateral lobe, respectively; pleural effusion will be M1a. With these alterations, cases staged as IB - T2b N0 M0 will be IIA, cases staged IIB - T2a N1 M0 will be IIA and cases IIIB- T4 N0- -1 M0 will be IIIA. The 7(th) TNM edition recommendations were applied to 203 broncho -pulmonary carcinomas, concerning epidermoid carcinomas (83) and adenocarcinomas (120) registered in the archive of the Serviço de Anatomia Patológica of the Hospitais da Universidade de Coimbra - Portugal, previously submitted to surgical resection and lymph node excision. The following alterations will be kept as the application of the future 7(th) TNM edition: 20 cases in stage IB will move to stage IIA (17) and stage IIB (3); 18 cases will change from stage IIB to stage IIA (17) and 1 case to stage IIIA; 2 cases from stage IIIB will move to stage IV; 6 cases in stage IV will move to stage IIIA (5) and 1 case to stage IIIB. In this translational adaptation from 6th to 7th TNM staging, 51 out of the 203 analysed cases change their staging, corresponding to 25.1%.

Padrão alimentar da dieta vegetariana / Eating pattern of vegetarian diet

Esta revisão reúne diversos estudos que avaliam os fatores que influenciam a adesão ao vegetarianismo, bem como o padrão alimentar e recomendações do guia vegetariano. Para tanto, foi realizado um levantamento bibliográfico mediante consulta às bases de dados Medline (National Library of Medicine, USA) e Lilacs (Bireme, Brasil), nas quais foram selecionadas publicações científicas em português e inglês, a partir dos seguintes termos: vegetariano, dieta vegetariana e vegetarianismo. Durante séculos o vegetarianismo tem sido difundido. Ao longo da história, grandes médicos, cientistas e filósofos seguiram a prática da alimentação vegetariana fundamentada em diferentes concepções. O crescente número de vegetarianos atribui aos profissionais da Saúde, sobretudo nutricionistas, a responsabilidade de conhecer os princípios da dieta vegetariana na expectativa de que, com base em evidências científicas, eles possam direcionar adequadamente sua conduta com respeito a essa dieta, bem como adequar esse padrão alimentar às necessidades nutricionais do indivíduo.