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Phytoplankton cells are now recognized as dynamic entities rather than as passive and isolated particles because they can actively modulate impacts of selection factors (nutrients, light, turbidity, and mixing) through a wide range of adaptations. Cell shape and/or chain length modulation is one of these processes but has predominantly been studied as an adaptation or an acclimatation to a specific growth limitation (light, nutrients, predation, etc.). In this study we have demonstrated that cell shape and size may have greater roles than previously known in phytoplankton ecology and species adaptation by permitting cell‐to‐cell signaling and more complex ecological processes that result from it. By exploring microscale biophysical interactions that lead to specific cell reorientation processes, we demonstrated that cell geometry not only modulates cell sinking rates but can also provide fast sensor responses to the cells' environment. Although gyrotaxis has been described in detail for motile phytoplankton cells, our findings illustrate that the reorientation process described here can occur even in non‐motile cells within their natural environment. An additional consistent behavior was also recently described for a diatom species (Pseudo‐nitzschia delicatessima), and with this study, we extend this observation to Pseudo‐nitzschia pungens and Pseudo‐nitzschia fraudulenta. Our observations emphasize the generality of this process, which adds a new level of complexity to our understanding of cellular interactions and their network of sensors. [ABSTRACT FROM AUTHOR]

Vaccination through the upper respiratory tract is a promising strategy, and particulate antigens, such as antigens associated with nanoparticles, triggered a stronger immune response than the sole antigens. Cationic maltodextrin-based nanoparticles loaded with phosphatidylglycerol (NPPG) are efficient for intranasal vaccination but non-specific to trigger immune cells. Here we focused on phosphatidylserine (PS) receptors, specifically expressed by immune cells including macrophages, to improve nanoparticle targeting through an efferocytosis-like mechanism. Consequently, the lipids associated with NPPG have been substituted by PS to generate cationic maltodextrin-based nanoparticles with dipalmitoyl-phosphatidylserine (NPPS). Both NPPS and NPPG exhibited similar physical characteristics and intracellular distribution in THP-1 macrophages. NPPS cell entry was faster and higher (two times more) than NPPG. Surprisingly, competition of PS receptors with phospho-L-serine did not alter NPPS cell entry and annexin V did not preferentially interact with NPPS. Although the protein association is similar, NPPS delivered more proteins than NPPG in cells. On the contrary, the proportion of mobile nanoparticles (50%), the movement speed of nanoparticles (3 µm/5 min), and protein degradation kinetics in THP-1 were not affected by lipid substitution. Together, the results indicate that NPPS enter cells and deliver protein better than NPPG, suggesting that modification of the lipids of cationic maltodextrin-based nanoparticles may be a useful strategy to enhance nanoparticle efficacy for mucosal vaccination.

Immunofluorescence on cleared intact cochlea allows detailed analysis of the cochlear ultrastructure, while avoiding the problems of dissection and serial sections. Protocols have been developed for mice and Mongolian gerbils. This technical note proposes a detailed and optimised immunofluorescence protocol in the Mongolian gerbil comprising significant quantitative and qualitative improvements. This protocol sequentially comprises: fixation (1 day), decalcification (6 days), pre-treatment (7.5 hours), immunolabelling (42 hours), dehydration and clearing (23 hours), followed by mounting and laser scanning confocal microscopy acquisition. This protocol has been optimised in terms of duration (10 days versus 13 days) with a reduction of the number of steps, improvement of the specificity of immunolabelling and optimisation of the quality of the results obtained. This technical note provides a detailed description of this protocol. [ABSTRACT FROM AUTHOR]

Les interactions entre le polynucléaire éosinophile et le mastocyte ont été décrites dans des maladies inflammatoires et/ou allergiques variées (Galdiero et al., 2017). L'éosinophilie médullaire est une caractéristique classique de la biopsie ostéomédullaire (BOM) de la mastocytose systémique, mais le rôle potentiel des éosinophiles n'a jamais été exploré. Nous avons analysé 61 BOM de patients avec mastocytoses indolentes (n = 33), « smouldering » (n = 4), ou avancées (n = 24), ou contrôles (n = 8). Des immunomarquages anti-EPX (éosinophiles), anti-c-KIT (mastocytes) ou des doubles immunomarquages EPX/c-KIT ont été réalisés en immunohistochimie (IHC) et en immunofluorescence (IF) avec microscopie confocale. Les surfaces d'immunomarquages c-KIT ou EPX sont exprimées en pourcentages par rapport aux surfaces cellulaires non marquées : par exemple, la surface d'immunomarquage EPX est calculée comme la surface des infiltrats cellulaires marqués EPX/(somme des surfaces EPX+ et des surfaces non marquées EPX). L'expression de marqueurs de surface par les mastocytes médullaires a été analysée en cytométrie en flux (CMF), et des images en IF ont été utilisées pour conforter l'analyse de l'expression du récepteur de l'IL-5. L'expression de marqueurs de surface par les éosinophiles sanguins a été analysée en CMF, et des analyses moléculaires par NGS ont été réalisées sur éosinophiles triés. En utilisant une analyse automatisée non supervisée des immunomarquages sur lames entières, nous avons observé une surface d'immunomarquage EPX, représentant à la fois les infiltrats d'éosinophiles intacts et les éosinophiles dégranulés, plus importante dans les mastocytoses non avancées (ISM + SSM) (médiane [IQR] : 37 % [19 ; 61]) que dans les contrôles (7 % [2,25 ; 17,25], p = 0,0003) et mastocytoses avancées (17 % [9,25 ; 36,5], p = 0,0014). Dans les mastocytoses non avancées, nous avons observé des corrélations positives entre les taux de tryptase sérique et le pourcentage d'éosinophiles au myélogramme (coefficient r de Spearman de 0,38, p = 0,038), le pourcentage d'éosinophiles sur les BOM (r 0,45, p = 0 0,007), la surface d'immunomarquage EPX (r 0,37, p = 0,035) et le niveau de dégranulation des éosinophiles (r 0,39, p = 0,023). Le nombre d'éosinophiles sur les BOM était également corrélé avec le nombre de mastocytes (r 0,47, p = 0,006) et la surface d'immunomarquage c-KIT sur les BOM (r 0,49, p = 0,003). Aucun de ces résultats n'était observé dans les mastocytoses avancées. Dans les mastocytoses non avancées, nous avons observé en IHC et en IF que les infiltrats d'éosinophiles et de larges plages de dégranulation étaient présents au sein et autour des infiltrats mastocytaires. En CMF, les mastocytes médullaires exprimaient à leur surface les récepteurs des interleukines (IL) 5 (confirmé en IF), IL-9 et IL-3, et les récepteurs de chimiokines PGD2, éotaxines et TARC, qui pourraient contribuer aux interactions réciproques avec les éosinophiles. De plus, les données de NGS suggèrent que les hyperéosinophilies sanguines ne soient pas d'origine clonale dans la mastocytoses non avancées : chez 4 patients avec mastocytose indolente et hyperéosinophilie > 1500/mm3, 1 patiente n'avait pas de mutation c-KIT, 3 patients étaient mutés D816 V, et la fréquence allélique était dans ce cas < 5 % dans les éosinophiles triés, sans autre mutation sur-représentée dans les éosinophiles par rapport à l'ADN médullaire total. Enfin, les éosinophiles sanguins analysés en CMF ont un profil d'activation caractérisé par une augmentation des intégrines CD44 et CD62L, et de CD137/4-1BB. Les nombreux médiateurs qui peuvent activer et/ou attirer les 2 cellules pourraient expliquer ces colocalisations et les corrélations observées entre des paramètres liés au mastocyte (tryptase sérique, nombre de mastocytes...) et à l'éosinophile (nombre de cellules, niveau de dégranulation...). Une efficacité potentielle des anti-IL-5 dans la mastocytose systémique indolente et le syndrome d'activation mastocytaire est d'ailleurs suggérée (De Wilde et al., 2016 ;Guillet et al., 2021). Nos données, confrontées aux propriétés connues des éosinophiles et des mastocytes, suggèrent que les deux populations cellulaires pourraient interagir dans les mastocytoses non avancées. Ainsi, les éosinophiles pourraient contribuer à la production de tryptase par les mastocytes et contribuer aux manifestations cliniques liées à l'activation mastocytaire. Des essais cliniques évaluant le bénéfice et la tolérance des anticorps monoclonaux anti-IL-5 ou anti-IL-5R semblent à envisager dans les mastocytoses non avancées. [ABSTRACT FROM AUTHOR]

• Better knowledge of the critical role of mitochondrion in FP and Q-UFP-induced lung toxicity. • Mitochondrial ROS overproduction and activation of NRF2-ARE signaling pathways. • Mitochondrial dynamics modifications in favor of accentuated fission process. • Mitochondrial quality control and metabolism dysfunctions as underlying mechanism of toxicity. • New insights into the physiopathology-induced by FP and mostly Q-UFP within the lung. Nowadays ambient particulate matter (PM) levels still regularly exceed the guideline values established by World Health Organization in most urban areas. Numerous experimental studies have already demonstrated the airway toxicity of the fine fraction of PM (FP), mainly triggered by oxidative stress-induced airway inflammation. However, only few studies have actually paid close attention to the ultrafine fraction of PM (UFP), which is likely to be more easily internalized in cells and more biologically reactive. Mitochondria are major endogenous sources of reactive oxygen species (ROS) through oxidative metabolism, and coordinate many critical cellular signaling processes. Mitochondria have been often studied in the context of PM toxicity and generally associated with apoptosis activation. However, little is known about the underlying adaptation mechanisms that could occur following exposure at sub-apoptotic doses of ambient PM. Here, normal human bronchial epithelial BEAS-2B cells were acutely or repeatedly exposed to relatively low doses (5 µg.cm−2) of FP (PM 2.5-0.18) or quasi-UFP (Q-UFP; PM 0.18) to better access the critical changes in mitochondrial morphology, functions, and dynamics. No significant cytotoxicity nor increase of apoptotic events were reported for any exposure. Mitochondrial membrane potential (ΔΨm) and intracellular ATP content were also not significantly impaired. After cell exposure to sub-apoptotic doses of FP and notably Q-UFP, oxidative phosphorylation was increased as well as mitochondrial mass, resulting in increased production of mitochondrial superoxide anion. Given this oxidative boost, the NRF2-ARE signaling pathway was significantly activated. However, mitochondrial dynamic alterations in favor of accentuated fission process were observed, in particular after Q-UFP vs FP, and repeated vs acute exposure. Taken together, these results supported mitochondrial quality control and metabolism dysfunction as an early lung underlying mechanism of toxicity, thereby leading to accumulation of defective mitochondria and enhanced endogenous ROS generation. Therefore, these features might play a key role in maintaining PM-induced oxidative stress and inflammation within lung cells, which could dramatically contribute to the exacerbation of inflammatory chronic lung diseases. The prospective findings of this work could also offer new insights into the physiopathology of lung toxicity, arguably initiate and/or exacerbate by acutely and rather repeated exposure to ambient FP and mostly Q-UFP. [ABSTRACT FROM AUTHOR]