BACKGROUND:This study is the first to evaluate a phenolic extract of Salvia officinalis in the TNBS murine model of colitis. Inflammatory bowel disease (IBD) involves transmural oxidative and nitrosative stress that up-regulates inducible nitric-oxide synthase (iNOS) and cyclo‑oxygenase-2 (COX-2). Although phenolic constituents of S. officinalis are anti-inflammatory in vitro, their efficacy has never been tested in the trinitrobenzene-sulphonic acid (TNBS) model of colitis. METHODS:Aqueous-ethanolic sage leaves were extracted and standardized [(total phenolics = 6208 ± 200 mg gallic acid equivalent (GAE) L-1; rosmarinic acid = 1.4 g/L). Male CD-1 mice received intracolonic TNBS (2.5 % w/v, 100 μL). Two hours later, and once daily for four days, they were gavaged with vehicle or sage extract (15 mg phenolic acids kg-1). Disease-activity index, colon length, macroscopic and histological scores, iNOS/COX-2 immunostaining, and survival (Kaplan-Meier) were recorded. RESULTS:Extract markedly attenuated TNBS injury: diarrhea score fell from 3.0 ± 0.0 to 0.3 ± 0.2, ulcer length from 3.6 ± 0.1 cm to 0.8 ± 0.2 cm, and colon shortening was prevented (11.8 ± 0.2 to 12.9 ± 0.2 cm). Mortality dropped from 36 % to 0 %. Histology improved (score 3 to 1) and iNOS, COX-2 over-expression was normalized (4.25 to 2.19-fold and 2.48 to 1.03-fold, respectively). Ancillary paw-edema and cell-migration assays confirmed anti-inflammatory activity without indicating anti-metastatic effects. CONCLUSIONS:The extract given orally affords the first demonstrated protection against acute TNBS colitis, normalizing key inflammatory markers and abolishing procedure-related mortality. Sage phenolics indicate promising multitarget leads for IBD therapy, meriting chronic-model and pharmacokinetic evaluation.

Multiple Sclerosis (MS) is the leading inflammatory and non-traumatic cause of disability in young adults, with late-onset MS emerging in middle-aged patients often resulting in poorer treatment responses and worse prognoses. The calcium-binding protein S100B is elevated in MS patients, and its targeting has shown promise in reducing disease severity in experimental autoimmune encephalomyelitis (EAE) models. However, most studies on MS pathology have focused on young animal models, leaving a gap in understanding the effects of age and S100B ablation on disease progression throughout the lifespan. This study aimed to characterize EAE in mice of different ages, examining demyelination, inflammation, and immune responses to determine whether S100B ablation could mitigate MS pathogenesis across the lifespan. EAE was induced in six cohorts of C57BL/6 mice: young adults (3 months), older adults (6 months), and middle-aged (12 months), including corresponding S100B knockout (KO) groups, followed for 23 days. Upon sacrifice, spinal cords were assessed via immunohistochemistry and Real-Time qPCR, while splenocytes were analyzed for immune cell characterization. Results indicated a more severe disease course in 12-month-old mice, marked by increased gliosis, inflammation, and impaired microglial phagocytic activity. Notably, S100B absence reduced gliosis and inflammatory markers across all ages, with 12-month-old S100B KO mice showing increased regulatory T cells. These findings highlight the exacerbating role of age and elevated S100B in MS progression, underscoring the importance of identifying age-specific MS markers and therapeutic targets.

Fluorescent molecules play a crucial role in biomedicine by facilitating the visualization and tracking of biological processes with sensitivity and specificity. However, tailoring their structure to meet the demands of live cell and in vivo imaging presents a significant challenge due to the intricate interplay of factors governing their structural and photophysical properties. In this study, we explored the potential of using multivariate linear free-energy relationships (mLFER) to optimize a multicomponent fluorescent platform. We prepared a small library of 20 fluorescent boronic-acid-derived salicylidenehydrazone (BASHY) complexes using a versatile reaction protocol and characterized their chemical stability in water-containing media. The obtained data served as input for the development of an mLFER model, enabling the prediction of a new BASHY dye and unraveling previously unknown mechanisms governing the stability of this unique platform of fluorescent dyes. The optimized dye was successfully employed in live cell experiments and in zebrafish larvae.

center dot Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a rare disorder caused by loss-of-function mutations in the sacsin chaperone. center dot Sacsin was initially described as a neuronal protein but is also found in astroglia and microglia. center dot We developed and characterized a new microglial cell model of ARSACS based on human HMC3 cells.. center dot HMC3Sacs- /- cells show aberrant distribution of vimentin and mitochondrial networks, and other alterations with potential consequences on neuroinflammation. center dot Our results support a possible role for microglial cells in ARSACS and provide a new tool to understand glial-specific mechanisms involved in this pathology

Chronic stress is a major risk factor for neuropsychiatric conditions such as depression. Adult hippocampal neurogenesis (AHN) has emerged as a promising target to counteract stress-related disorders given the ability of newborn neurons to facilitate endogenous plasticity. Recent data sheds light on the interaction between cannabinoids and neurotrophic factors underlying the regulation of AHN, with important effects on cognitive plasticity and emotional flexibility. Since physical exercise (PE) is known to enhance neurotrophic factor levels, we hypothesised that PE could engage with cannabinoids to influence AHN and that this would result in beneficial effects under stressful conditions. We therefore investigated the actions of modulating cannabinoid type 2 receptors (CB2R), which are devoid of psychotropic effects, in combination with PE in chronically stressed animals. We found that CB2R inhibition, but not CB2R activation, in combination with PE significantly ameliorated stress-evoked emotional changes and cognitive deficits. Importantly, this combined strategy critically shaped stress-induced changes in AHN dynamics, leading to a significant increase in the rates of cell proliferation and differentiation of newborn neurons, overall reduction in neuroinflammation, and increased hippocampal levels of BDNF. Together, these results show that CB2Rs are crucial regulators of the beneficial effects of PE in countering the effects of chronic stress. Our work emphasises the importance of understanding the mechanisms behind the actions of cannabinoids and PE and provides a framework for future therapeutic strategies to treat stress-related disorders that capitalise on lifestyle interventions complemented with endocannabinoid pharmacomodulation.

Recent advances in understanding Alzheimer's disease (AD) suggest the possibility of an infectious etiology, with Porphyromonas gingivalis emerging as a prime suspect in contributing to AD. P. gingivalis may invade systemic circulation via weakened oral/intestinal barriers and then cross the blood-brain barrier (BBB), reaching the brain and precipitating AD pathology. Based on the proposed links between P. gingivalis and AD, a prospective approach is the development of an oral nanovaccine containing P. gingivalis antigens for mucosal delivery. Targeting the gut-associated lymphoid tissue (GALT), the nanovaccine may elicit both mucosal and systemic immunity, thereby hampering P. gingivalis ability to breach the oral/intestinal barriers and the BBB, respectively.The present study describes the optimization, characterization, and in vitro evaluation of a candidate chitosan-coated poly(lactic-co-glycolic acid) (PLGA-CS) nanovaccine containing a P. gingivalis antigen extract. The nanocarrier was prepared using the double emulsion solvent evaporation method and optimized for selected experimental factors, e.g. PLGA amount, surfactant concentration, w1/o phase ratio, applying a D-optimal statistical design to target the desired physicochemical criteria for its intended application. After nanocarrier optimization, the nanovaccine was characterized in terms of particle size, polydispersity index (PdI), ζ-potential, encapsulation efficiency (EE), drug loading (DL), morphology, and in vitro release profile, as well as for mucoadhesivity, stability under simulated gastrointestinal conditions, antigen integrity, in vitro cytotoxicity and uptake using THP-1 macrophages.The candidate PLGA-CS nanovaccine demonstrated appropriate physicochemical, mucoadhesive, and antigen release properties for oral delivery, along with acceptable levels of EE (55.3 ± 3.5%) and DL (1.84 ± 0.12%). The integrity of the encapsulated antigens remained uncompromised throughout NPs production and simulated gastrointestinal exposure, as confirmed by SDS-PAGE and Western blotting analyses. Furthermore, the nanovaccine showed effective in vitro uptake, while exhibiting low cytotoxicity. Taken together, these findings underscore the potential of PLGA-CS NPs as carriers for adequate antigen mucosal delivery, paving the way for further investigations into their applicability as vaccine candidates against P. gingivalis.

Multiple Sclerosis (MS), a neuroinflammatory disease of the central nervous system, is one of the commonest causes of non-traumatic disability among young adults. Impaired cognition arises as an impactful symptom affecting more than 50% of the patients and with substantial impact on social, economic, and individual wellbeing. Despite the lack of therapeutic strategies, many efforts have been made to understand the mechanisms behind cognitive impairment in MS patients. Here, we aimed to investigate whether microglia-derived synaptic elimination and immune interactions are exacerbated in MS patients with impaired cognition when compared to non-demented controls (NDC) and cognitively preserved MS patients, that may clarify the role of immune cell interplay in MS cognitive deficits. Postmortem hippocampal samples were obtained from NDCs and MS patients. Sixteen MS patients were categorized based on their cognitive status: preserved cognition (MSCP) and impaired cognition (MSCI). Immunohistochemistry studies were conducted to explore the density of microglia, their role in synaptic engulfment, and their interaction with CD8+ immune cells in the context of cognitive impairment in MS. In high synaptic density hippocampal regions, MSCI patients exhibited a massive presence of microglia cells actively engulfing both excitatory and inhibitory synapses, accompanied by morphological alterations. Additionally, there was an increased expression of the complement protein C1q particularly localized at inhibitory synapses within microglia cells, suggesting a preferential engulfment of complement-tagged inhibitory synapses in MSCI patients. Furthermore, in hippocampal lesions of MSCI patients, we detected a significant infiltration of microglia and CD8 T cells that may be contributing to the smouldering MS and cognitive deterioration. These findings demonstrate that cognitive deficits occurring in MS are associated with microglia engulfment of C1q-tagged inhibitory synapses, which may be driven by direct or indirect stimulation from CD8+ T cells.

Oligodendrocytes (OLs) are vital for myelin formation in the Central Nervous System. OLs can be produced by the maturation of oligodendrocyte precursor cells (OPCs) present throughout the brain parenchyma or from the differentiation of subventricular zone-derived neural stem cells (SVZ-NSCs). Importantly, efficient differentiation from SVZ-NSCs remains a significant area of research due to its potential for remyelination in demyelinating disorders. In this work, we studied the role of brain-derived neurotrophic factor (BDNF) and adenosine A2A receptors (A2ARs), as well as the putative crosstalk between these two modulatory mechanisms, in regulating oligodendrogenesis from SVZ-NSCs. Using a neurosphere culture system, we observed that BDNF significantly increased the mRNA expression levels of OPC cell markers after 2 days in vitro (DIV), an effect blocked by the A2AR antagonist ZM 241385. This early transcriptional regulation by BDNF was followed by changes in the percentage of both OPCs and mature OLs in culture at DIV 7 and 14. Interestingly, blocking A2ARs prevented the potentiating effect of BDNF on the percentage of OLs at DIV 14. Concerning the morphology of mature OLs, BDNF influenced their maturation by reducing branching near the soma at DIV 7, an effect that was not observed at 14 DIV, when all treatments resulted in similar OL morphology. Overall, our results establish BDNF as a regulator of OL formation from SVZ-NSCs, with A2AR-BDNF interaction modulating the differentiation process. ### Competing Interest Statement The authors have declared no competing interest.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is a developmental and degenerative disorder caused by loss-of-function mutations in the gene that codifies for the sacsin chaperone. Sacsin was initially described as a neuronal protein but is found in various cell types, including astroglial, microglial, kidney, and skin cell lines. We and others have shown that virtually all cell and animal models of ARSACS show disruption of intermediate filament (IF) cytoskeleton and organelle distribution. This article extends previous observations from our lab on the C6 astroglial-like model and describes the development and characterization of a new human microglial cell model based on HMC3 cells. HMC3 cells knocked out for sacsin show similar alterations to C6 astroglial-like cells: aberrant distribution of IF network and organelles, downregulation of developmental transcription factors STAT3 and Smad1, and alterations in autophagy machinery and markers of intracellular stress, resulting in morphological changes. Our results extend previous observations suggesting a possible role for glial cells in ARSACS and provide new tools to understand the glial-specific mechanisms involved in this pathology. ### Competing Interest Statement The authors have declared no competing interest.

Ethylene removal is crucial for fruits and vegetables preservation because even a very low concentration (<0.1 ppm) can induce ripening during storage and transportation. Ag+ exchanged ZSM-5 zeolites were investigated as adsorbents for the efficient removal of ethylene. Two ZSM-5 materials (Si/Al ratios of 15 and 40), with different amounts of Ag (up to 6 % by weight) and two compensating cations, Na+ and H+, were used for this purpose. The adsorbents were characterized by X-ray diffraction, diffuse reflectance spectroscopy and H2-TPR experiments. Their performance in ethylene adsorption was determined by the means of breakthrough curves experiments, mimicking the atmospheric conditions of industrial fruits cold storage chambers, particularly the high relative humidity levels. Results show that adsorbents are highly efficient in removing ethylene (max. 500 mu mol g(-1) in the absence of water). Monovalent Ag+ species have been identified as the main contributors to the excellent performance of the different adsorbents, as they can easily interact with ethylene through strong pi interaction. Moreover, statistical analysis (ANOVA) results confirmed that, in the presence of water, Ag-based ZSM-5 materials with a higher Si/Al ratio and Na+ as the charge balancing cation, i.e., those with a higher hydrophobic character, are the best adsorbents for ethylene removal.

Obesity is a global epidemic, affecting roughly 30% of the world’s population and predicted to rise. This disease results from genetic, behavioral, societal, and environmental factors, leading to excessive fat accumulation, due to insufficient energy expenditure. The adipose tissue, once seen as a simple storage depot, is now recognized as a complex organ with various functions, including hormone regulation and modulation of metabolism, inflammation, and homeostasis. Obesity is associated with a low-grade inflammatory state and has been linked to neurodegenerative diseases like multiple sclerosis (MS), Alzheimer’s (AD), and Parkinson’s (PD). Mechanistically, reduced adipose expandability leads to hypertrophic adipocytes, triggering inflammation, insulin and leptin resistance, blood-brain barrier disruption, altered brain metabolism, neuronal inflammation, brain atrophy, and cognitive decline. Obesity impacts neurodegenerative disorders through shared underlying mechanisms, underscoring its potential as a modifiable risk factor for these diseases. Nevertheless, further research is needed to fully grasp the intricate connections between obesity and neurodegeneration. Collaborative efforts in this field hold promise for innovative strategies to address this complex relationship and develop effective prevention and treatment methods, which also includes specific diets and physical activities, ultimately improving quality of life and health.

Sixty percent of melanoma patients present brain metastases, having low response rates to current systemic therapies, which do not prolong survival. New therapeutic options to address this clinical need are urgent. Here, we report the development of a polymeric nanoparticle as a cancer nanovaccine capable of selectively co-targeting toll-like receptor function and PD-L1 expression to improve antigen presentation and subsequent effector immune cell function within brain microenvironment. Mannose-poly(lactic-co-glycolic acid)/poly(lactic acid) (man-PLGA/PLA) nanoparticles (NP) were prepared by a modified double emulsion solvent evaporation method, to deliver combinations of melanoma neoantigens, toll-like receptor ligands and regulators of the PD-1/PD-L1 axis. NP physicochemical properties were fully characterized, including size, surface charge and morphology. The amount of melanoma neoantigens and immune regulators entrapped within NP was determined by HPLC. Immature dendritic cells (DC) were used to evaluate the impact of NP on cell viability, and to assess NP uptake kinetics by flow cytometry. NP ability to target and trigger the activation of DC and T cells, was assessed in the lymph nodes and spleen of immunized mice. The NP anti-tumor immune-mediated effect was evaluated in vivo in two primary melanoma-bearing immunocompetent mouse models (B16F10 and B16MO5), and in a melanoma brain metastasis (MBM) mouse model (B16F10), which also included the immune profiling within tumor site by flow cytometry, before and after treatment.NP presented an average diameter of 180 nm, narrow polydispersity index, surface charge close to neutrality, spherical morphology, and high loadings of the neoantigens and immunoregulators. NP did not affect DC viability and were extensively internalized by immature DC. NP were preferentially taken-up in vivo by DC, increasing the expression of activation/maturation markers at DC surface, such as CD80 and CD86. NP elicited antigen-specific T-cell responses, by the significant increase in the expression of TNF-alpha and IFN-gamma (TH1-guided response). In vivo combination of NP with anti-PD-L1 induced a potent immune-mediated anti-tumor response in both primary and preclinical models of MBM, overcoming tumor development with an increased overall survival. This combination re-shaped immune and stroma cell populations in primary tumor and MBM microenvironment, which presented a marked infiltration of cytotoxic T cells that correlated with a decreased expression of PD-1 and PD-L1 within the TME. The synergy between our nanovaccine and anti-PD-L1 provide essential insights to devise alternative combinations regiments to improve the efficacy of immune checkpoint inhibitors in metastatic melanoma, thus opening new line for this unmet medical need. Citation Format: Barbara Carreira, Rita A. Acúrcio, Sabina Pozzi, Ron Kleiner, Liane I. Moura, Ana I. Matos, Daniela vaskovich, Carina Peres, Adelaide Fernandes, Sara Xapelli, Ronit Satchi-Fainaro, Helena F. Florindo. Translational nanotechnology-based cancer vaccine to re-educate host immune response against metastatic cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 835.

History is full of women who made enormous contributions to science. While there is little to no imbalance at the early career stage, a decreasing proportion of women is found as seniority increases. In the multiple sclerosis (MS) field, 44% of first authors and only 35% of senior authors were female. So, in this review, we highlight ground-breaking research done by women in the field of MS, focusing mostly on their work as principal investigators. MS is an autoimmune disorder of the central nervous system (CNS), with evident paradigm shifts in the understating of its pathophysiology. It is known that the immune system becomes overactivated and attacks myelin sheath surrounding axons. The resulting demyelination disrupts the communication signals to and from the CNS, which causes unpredictable symptoms, depending on the neurons that are affected. Classically, MS was reported to cause mostly physical and motor disabilities. However, it is now recognized that cognitive impairment affects more than 50% of the MS patients. Another shifting paradigm was the involvement of gray matter in MS pathology, formerly considered to be a white matter disease. Additionally, the identification of different T cell immune subsets and the mechanisms underlying the involvement of B cells and peripheral macrophages provided a better understanding of the immunopathophysiological processes present in MS. Relevantly, the gut-brain axis, recognized as a bi-directional communication system between the CNS and the gut, was found to be crucial in MS. Indeed, gut microbiota influences not only different susceptibilities to MS pathology, but it can also be modulated in order to positively act in MS course. Also, after the identification of the first microRNA in 1993, the role of microRNAs has been investigated in MS, either as potential biomarkers or therapeutic agents. Finally, concerning MS therapeutical approaches, remyelination-based studies have arisen on the spotlight aiming to repair myelin loss/neuronal connectivity. Altogether, here we emphasize the new insights of remarkable women that have voiced the impact of cognitive impairment, white and gray matter pathology, immune response, and that of the CNS-peripheral interplay on MS diagnosis, progression, and/or therapy efficacy, leading to huge breakthroughs in the MS field.

The experimental autoimmune encephalomyelitis (EAE) model is the most commonly used animal model of multiple sclerosis (MS). However, phenotypic characterization of mice based on the traditional 5-point clinical paralysis scale does not fully capture disease progression. The frailty index (FI) conceptualizes frailty as the accumulation of health deficits and it is widely used to assess overall health in aging humans and preclinical models. Here, we adapted an established mouse FI tool for use in EAE mice and determined whether this could evaluate general signs of health in variably aged female EAE mice. The EAE-Clinical FI included 34 items related to clinical signs and deficits characteristic of aging and MS. This tool clearly showed more detailed EAE progression and severity at all ages, highlighting changes in systems other than motor paralysis measured with the traditional 5-point paralysis scale. When we induced disease at 3 and 6 months of age, mice showed typical EAE clinical manifestations with peak disease severity between 17 and 19 days post-induction and mean frailty scores of 0.36 ± 0.04 (3-month-old) and 0.43 ± 0.05 (6-month-old). By contrast, disease severity peaked after 14 days in 12-month-old mice. They showed atypical signs including wobbling, early belly drag, and splayed hindlegs that were better captured with the EAE-Clinical FI. Peak frailty scores also were higher than those of younger animals (0.54 ± 0.04). As MS most often develops in young to middle-aged people, this new tool may have significant value for use in EAE animal studies as a first step toward translation to people with MS.