Patient-derived follicular lymphoma spheroids recapitulate lymph node signaling and immune profile uncovering galectin-9 as a novel immunotherapeutic target.

Follicular lymphoma (FL), the most common indolent non-Hodgkin lymphoma, constitutes a paradigm of immune tumor microenvironment (TME) contribution to disease onset, progression, and heterogenous clinical outcome. Here we present the first FL-Patient Derived Lymphoma Spheroid (FL-PDLS), including fundamental immune actors and features of TME in FL lymph nodes (LNs). FL-PDLS is organized in disc-shaped 3D structures composed of proliferating B and T cells, together with macrophages with an intermediate M1/M2 phenotype. FL-PDLS recapitulates the most relevant B-cell transcriptional pathways present in FL-LN (proliferation, epigenetic regulation, mTOR, adaptive immune system, among others). The T cell compartment in the FL-PDLS preserves CD4 subsets (follicular helper, regulatory, and follicular regulatory), also encompassing the spectrum of activation/exhaustion phenotypes in CD4 and CD8 populations. Moreover, this system is suitable for chemo and immunotherapy testing, recapitulating results obtained in the clinic. FL-PDLS allowed uncovering that soluble galectin-9 limits rituximab, rituximab, plus nivolumab/TIM-3 antitumoral activities. Blocking galectin-9 improves rituximab efficacy, highlighting galectin-9 as a novel immunotherapeutic target in FL. In conclusion, FL-PDLS maintains the crosstalk between malignant B cells and the immune LN-TME and constitutes a robust and multiplexed pre-clinical tool to perform drug screening in a patient-derived system, advancing toward personalized therapeutic approaches.

Patient-derived lymphoma spheroids integrating immune tumor microenvironment as preclinical follicular lymphoma models for personalized medicine.

BACKGROUND: Follicular lymphoma (FL), the most common indolent non-Hodgkin's Lymphoma, is a heterogeneous disease and a paradigm of the contribution of immune tumor microenvironment to disease onset, progression, and therapy resistance. Patient-derived models are scarce and fail to reproduce immune phenotypes and therapeutic responses. METHODS: To capture disease heterogeneity and microenvironment cues, we developed a patient-derived lymphoma spheroid (FL-PDLS) model culturing FL cells from lymph nodes (LN) with an optimized cytokine cocktail that mimics LN stimuli and maintains tumor cell viability. RESULTS: FL-PDLS, mainly composed of tumor B cells (60% on average) and autologous T cells (13% CD4 and 3% CD8 on average, respectively), rapidly organizes into patient-specific three-dimensional (3D) structures of three different morphotypes according to 3D imaging analysis. RNAseq analysis indicates that FL-PDLS reproduces FL hallmarks with the overexpression of cell cycle, BCR, or mTOR signaling related gene sets. FL-PDLS also recapitulates the exhausted immune phenotype typical of FL-LN, including expression of BTLA, TIGIT, PD-1, TIM-3, CD39 and CD73 on CD3+ T cells. These features render FL-PDLS an amenable system for immunotherapy testing. With this aim, we demonstrate that the combination of obinutuzumab (anti-CD20) and nivolumab (anti-PD1) reduces tumor load in a significant proportion of FL-PDLS. Interestingly, B cell depletion inversely correlates with the percentage of CD8+ cells positive for PD-1 and TIM-3. CONCLUSIONS: In summary, FL-PDLS is a robust patient-derived 3D system that can be used as a tool to mimic FL pathology and to test novel immunotherapeutic approaches in a context of personalized medicine.

γδ T cells in immunotherapies for B-cell malignancies.

Despite the advancements in therapy for B cell malignancies and the increase in long-term survival of patients, almost half of them lead to relapse. Combinations of chemotherapy and monoclonal antibodies such as anti-CD20 leads to mixed outcomes. Recent developments in immune cell-based therapies are showing many encouraging results. γδ T cells, with their potential of functional plasticity and their anti-tumoral properties, emerged as good candidates for cancer immunotherapies. The representation and the diversity of γδ T cells in tissues and in the blood, in physiological conditions or in B-cell malignancies such as B cell lymphoma, chronic lymphoblastic leukemia or multiple myeloma, provides the possibility to manipulate them with immunotherapeutic approaches for these patients. In this review, we summarized several strategies based on the activation and tumor-targeting of γδ T cells, optimization of expansion protocols, and development of gene-modified γδ T cells, using combinations of antibodies and therapeutic drugs and adoptive cell therapy with autologous or allogenic γδ T cells following potential genetic modifications.

A novel patient-derived 3D model recapitulates mantle cell lymphoma lymph node signaling, immune profile and in vivo ibrutinib responses.

Mantle cell lymphoma (MCL), a rare and aggressive B-cell non-Hodgkin lymphoma, mainly develops in the lymph node (LN) and creates a protective and immunosuppressive niche that facilitates tumor survival, proliferation and chemoresistance. To capture disease heterogeneity and tumor microenvironment (TME) cues, we have developed the first patient-derived MCL spheroids (MCL-PDLS) that recapitulate tumor oncogenic pathways and immune microenvironment in a multiplexed system that allows easy drug screening, including immunotherapies. MCL spheroids, integrated by tumor B cells, monocytes and autologous T-cells self-organize in disc-shaped structures, where B and T-cells maintain viability and proliferate, and monocytes differentiate into M2-like macrophages. RNA-seq analysis demonstrated that tumor cells recapitulate hallmarks of MCL-LN (proliferation, NF-kB and BCR), with T cells exhibiting an exhaustion profile (PD1, TIM-3 and TIGIT). MCL-PDLS reproduces in vivo responses to ibrutinib and demonstrates that combination of ibrutinib with nivolumab (anti-PD1) may be effective in ibrutinib-resistant cases by engaging an immune response with increased interferon gamma and granzyme B release. In conclusion, MCL-PDLS recapitulates specific MCL-LN features and in vivo responses to ibrutinib, representing a robust tool to study MCL interaction with the immune TME and to perform drug screening in a patient-derived system, advancing toward personalized therapeutic approaches.

3D Model Characterization by 2D and 3D Imaging in t(14;18)-Positive B-NHL: Perspectives for In Vitro Drug Screens in Follicular Lymphoma.

Follicular lymphoma (FL) is an indolent B cell lymphoproliferative disorder of transformed follicular center B cells, which accounts for 20-30 percent of all non-Hodgkin lymphoma (NHL) cases. Great advances have been made to identify the most relevant targets for precision therapy. However, no relevant models for in vitro studies have been developed or characterized in depth. To this purpose, we generated a 3D cell model from t(14;18)-positive B-NHL cell lines cultured in ultra-low attachment 96-well plates. Morphological features and cell growth behavior were evaluated by classical microscopy (2D imaging) and response to treatment with different drugs was evaluated by a high-content analysis system to determine the robustness of the model. We show that the ultra-low attachment (ULA) method allows the development of regular, spherical and viable ULA-multicellular aggregates of lymphoma cells (MALC). However, discrepancies in the results obtained after 2D imaging analyses on drug-treated ULA-MALC prompted us to develop 3D imaging and specific analyses. We show by using light sheet microscopy and specifically developed 3D imaging algorithms that 3D imaging and dedicated analyses are necessary to characterize morphological properties of 3D models and drug effects. This study proposes a new method, but also imaging tools and informatic solutions, developed for FL necessary for future preclinical studies.

Participação em programas de intervenção comunitária e qualidade de vida: resultados de um estudo multicêntrico em Portugal / Participation in community intervention programmes and quality of life: findings from a multicenter study in Portugal

Objetivo: Analisar a qualidade de vida (QV) em indivíduos que participam de programas de intervenção comunitária (PIC) orientados para uma vida ativa e saudável. Método: Estudo transversal multicêntrico com 304 participantes, com 55 anos ou mais de idade, a viver na comunidade em três localidades portuguesas. Metade desses indivíduos (n=152) envolvida em PIC (grupo de intervenção). Esse grupo foi emparelhado segundo sexo e grupo etário com número equivalente de participantes (n=152) que não frequenta PIC (grupo de comparação). As atividades dos PIC foram agrupadas segundo a sua natureza: sociorrecreativas, educativas/aprendizagem ao longo da vida (ALV) e atividade física. Recolheu-se informação usando Questionário de Participação Social, WHOQOL-Bref e Escala de Satisfação com a Vida. Resultados: Os participantes dos PIC tinham média de idade de 71,4 (±5,4) anos, eram predominantemente mulheres (75,0%), casados (65,4%), com escolaridade inferior a cinco anos (71,7%) e rendimento familiar mensal até 750 euros (47,4%). O GI apresentou melhor QV no domínio físico do que o GC ( p<0,03). A atividade física foi a modalidade mais frequentada nos PIC (n=119; 78,3%) em comparação com atividades educativas/ALV (n=46; 30,3%) e sociorrecreativas (n=25; 16,4%). Os praticantes de atividade física em PIC apresentaram melhor QV nos domínios psicológico, relações sociais e ambiente do que os não praticantes ( p<0,05). Conclusão: A participação em PIC está associada à QV pelo que, em linha com o quadro do envelhecimento ativo, se recomenda implementar PIC no âmbito das políticas públicas, promovendo sistematicamente a QV da população. AU

Objective: The present study aimed to analyze quality of life (QoL) in participants of community intervention programs (CIP) focused on healthy aging. Method: A multicenter cross-sectional study was carried out with 304 community-dwelling participants, aged 55 years old or more and living in three locations in Portugal. Half of these individuals (n=152) were involved in a CIP (intervention group). The intervention group was paired according to sex and age group with an equivalent number of participants (n=152) that did not take part in a CIP (comparison group). Activities implemented in the CIP were grouped according to their nature: socio-recreational, educational/lifelong learning and physical activity. Data collection involved a Social Participation Questionnaire, the WHOQOL-Bref and the Satisfaction With Life Scale. Results: The CIP participants (n=152) had a mean age of 71.4 years (±5.4), were predominantly women (75.0%), married (65.4%), with fewer than five years of education (71.7%) and a monthly family income of up to 750 euros (47.4%). The intervention group had a significantly higher QoL in the physical domain than the comparison group ( p<0.03). Physical activity was the most frequently attended session in the CIP (n=119, 78.3%), in comparison with educational/ lifelong learning (n=46, 30.3%) and socio-recreational (n=25, 16.4%) activities. People practicing physical activity in the CIP had a significantly higher QoL in the psychological, social relationships and environment domains ( p<0.05). Conclusion: Participation in the CIP was associated with QoL. Therefore, in line with the active aging framework, CIPs must be a part of public policy measures aimed at the QoL of the population.AU

Occurrence of toxigenic Aspergillus flavus in commercial Bulgur wheat

Abstract Aflatoxins are mutagenic, carcinogenic, and teratogenic mycotoxins. The objective of this work was to study the presence of aflatoxigenic Aspergillus in commercial Bulgur wheat in the city of Maringá, Paraná, Brazil. Thirty samples of commercial Bulgur wheat, acquired in the period of August 2011 to January 2012, were evaluated. The enumeration analysis showed that samples had up to 273.3 CFU of molds and 133.3 CFU of aflatoxigenic Aspergillus per gram of wheat. Forty-two monosporic isolates were obtained and identified as Aspergillus flavus. The isolates were analyzed regarding their aflatoxigenic potential by culture in coconut milk agar; hydroxide vapor exposure; chromatography; and polymerase chain reaction (PCR) targeting genes that code enzymes of the aflatoxins synthesis pathway. Some of the isolates were confirmed to be aflatoxin producers and several of them presented a genetic profile of aflatoxin synthesis. The obtained results demonstrated that Bulgur wheat A. flavus contamination is concerning.

Skeletal muscle regeneration and impact of aging and nutrition.

After skeletal muscle injury a regeneration process takes place to repair muscle. Skeletal muscle recovery is a highly coordinated process involving cross-talk between immune and muscle cells. It is well known that the physiological activities of both immune cells and muscle stem cells decline with advancing age, thereby blunting the capacity of skeletal muscle to regenerate. The age-related reduction in muscle repair efficiency contributes to the development of sarcopenia, one of the most important factors of disability in elderly people. Preserving muscle regeneration capacity may slow the development of this syndrome. In this context, nutrition has drawn much attention: studies have demonstrated that nutrients such as amino acids, n-3 polyunsaturated fatty acids, polyphenols and vitamin D can improve skeletal muscle regeneration by targeting key functions of immune cells, muscle cells or both. Here we review the process of skeletal muscle regeneration with a special focus on the cross-talk between immune and muscle cells. We address the effect of aging on immune and skeletal muscle cells involved in muscle regeneration. Finally, the mechanisms of nutrient action on muscle regeneration are described, showing that quality of nutrition may help to preserve the capacity for skeletal muscle regeneration with age.

Transição e (In)Adaptação ao Lar de Idosos: Um Estudo Qualitativo / Transition and (In)Adaptation to Nursing Home: A Qualitative Study

RESUMO Um dos contextos de envelhecimento é o lar de idosos, mas pouca atenção tem sido dedicada ao processo de institucionalização, particularmente numa perspectiva desenvolvimental e ecológica. Assim, desenhamos um estudo qualitativo com o objectivo de explorar a experiência de transição e adaptação ao lar de 15 idosos institucionalizados há mais de um ano. Os participantes foram entrevistados e a análise de conteúdo permitiu identificar um tema comum - Vivência da Institucionalização, que integra três domínios: Tomada de decisão, Processo de institucionalização, e Posicionamento face à institucionalização. Os resultados sugerem que a institucionalização constitui uma transição que nem sempre resulta em adaptação e que o envolvimento do idoso na tomada de decisão e as características ambientais são fundamentais neste processo.

ABSTRACT One of the aging contexts is the nursing home, but little attention has been dedicated to the institutionalization process, particularly from a developmental and ecological perspective. Thus, we designed a qualitative study to explore the experience of transition and adaptation of 15 elderly who were institutionalized in a nursing home for more than a year. Participants were interviewed and content analysis identified a common theme - Experience of Institutionalization, which integrates three domains: Decision making, Process of institutionalization, and Positioning to institutionalization. The results suggest that institutionalization is a transition that does not always result in adaptation, and that the elderly participation in the decision making and the environmental characteristics are fundamental in this process.

Versão portuguesa da escala de ansiedade filial / Portuguese version of the filial anxiety scale

O aumento da longevidade e do envelhecimento da população aumenta a probabilidade dos filhos adultos assumirem os cuidados dos seus pais envelhecidos. Quando os filhos percepcionam que os pais podem vir a necessitar de cuidados podem evidenciar uma preocupação antecipada relativamente à sua capacidade para cuidar e ao bem-estar dos mesmos. Cicirelli (1988) denominou esta preocupação de ansiedade filial e desenvolveu a Escala de Ansiedade Filial para a avaliar. O presente estudo tem como objectivo apresentar o processo de validação da Escala de Ansiedade Filial (EAF, Cicirelli, 1988) para a língua portuguesa (Europa). Participaram no estudo 130 filhos adultos de meia-idade portugueses. Os resultados sugerem que a versão portuguesa apresenta características psicométricas adequadas, mostrando-se um instrumento válido.

The growth of longevity and aging increase the likelihood of adult children provide care to their aging parents. However, adult children may exhibit an anticipatory worry about their proficiency to provide care to their parents as well as about parents’ well-being. Cicirelli (1988) labeled this worry as filial anxiety and developed the Filial Anxiety Scale to measure it. The aim of the current study is to describe the translation process and validation of the European Portuguese version of the Escala de Ansiedade Filial (EAF; Cicirelli, 1998), by reporting the EAF main psychometric properties in a sample of 130 Portuguese middle-aged adult children. By demonstrating the satisfactory psychometric properties of the EAF, the results suggest the EAF as a valid instrument to measure the filial anxiety.

Identification of new galactose oxidase genes in Fusarium spp.

Galactose oxidase (GO) converts galactose to an aldehyde and has several biotechnological applications, including cancer diagnosis. It is mainly produced by Fusarium austroamericanum but is also produced by Fusarium acuminatum and by isolates of the Fusarium graminearum and Gibberella fujikuroi complexes. The F. austroamericanum GO gaoA gene has been cloned, but the GO genes from other secreting species have not been characterized. Problems associated with the F. austroamericanum GO such as high pI and low catalytic efficiency and thermostability, and the difficult purification process makes the search for homologous genes attractive. In this work, the GO genes from Fusarium verticillioides and Fusarium subglutinans, two species of the G. fujikuroi complex, were cloned, sequenced, and analyzed. New GO genes were found in databases and were used to construct a phylogenetic tree, which revealed the existence of three orthologous lineages of GO genes in Fusarium spp. In addition, RT-PCR analyses revealed that the new GO cloned gene may be endogenously expressed in F. subglutinans but not in F. verticillioides, in the used culture conditions.