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Cognitive decline is a common feature of aging, particularly in memory domains supported by the medial temporal lobe (MTL). The ability to identify intervention strategies to treat or prevent this decline is challenging due to substantial variability between adults in terms of age of onset, rate and severity of decline, and many factors that could influence cognitive reserve. These factors can be somewhat mitigated by use of within-subject designs. Aged outbred Long-Evans rats have proven useful for identifying translationally relevant substrates contributing to age-related decline in MTL-dependent memory. In this population, some animals show reliable impairment on MTL-dependent tasks while others perform within the range of young adult rats. However, currently there are relatively few within-subject behavior protocols for assessing MTL function over time, and most require extensive training and appetitive motivation for associative learning. In the current study, we aimed to test whether water maze learning impairments in aged Long-Evans rats would be predictive of delayed recognition memory impairments and whether these odor memory impairments would be stable within subjects over multiple rounds of testing.

BackgroundImmunotherapy of cancer is an emerging field with the potential to improve long-term survival. Thus far, adoptive transfer of tumor-specific T cells represents an effective treatment option for tumors of the hematological system such as lymphoma, leukemia or myeloma. However, in solid tumors, treatment efficacy is low owing to the immunosuppressive microenvironment, on-target/off-tumor toxicity, limited extravasation out of the blood vessel, or ineffective trafficking of T cells into the tumor region. Superparamagnetic iron oxide nanoparticles (SPIONs) can make cells magnetically controllable for the site-specific enrichment. MethodsIn this study, we investigated the influence of SPION-loading on primary human T cells for the magnetically targeted adoptive T cell therapy. For this, we analyzed cellular mechanics and the T cell response after stimulation via an exogenous T cell receptor (TCR) specific for the melanoma antigen MelanA or the endogenous TCR specific for the cytomegalovirus antigen pp65 and compared them to T cells that had not received SPIONs.ResultsSPION-loading of human T cells showed no influence on cellular mechanics, therefore retaining their ability to deform to external pressure. Additionally, SPION-loading did not impair the T cell proliferation, expression of activation markers, cytokine secretion, and tumor cell killing after antigen-specific activation mediated by the TCR. ConclusionIn summary, we demonstrated that SPION-loading of T cells did not affect cellular mechanics or the functionality of the endogenous or an exogenous TCR, which allows future approaches using SPIONs for the magnetically enrichment of T cells in solid tumors.

Key pointsThe amount of fibrotic material in dystrophic mouse muscles relates to contractile function, but not passive function. Collagen fibres in skeletal muscle are associated with increased passive muscle stiffness in fibrotic muscles. The alignment of collagen is independently associated with passive stiffness in dystrophic skeletal muscles. These outcomes demonstrate that collagen architecture rather than collagen content should be a target of anti-fibrotic therapies to treat muscle stiffness.AbstractFibrosis is prominent in many skeletal muscle pathologies including dystrophies, neurological disorders, cachexia, chronic kidney disease, sarcopenia and metabolic disorders. Fibrosis in muscle is associated with decreased contractile forces and increased passive stiffness that limits joint mobility leading to contractures. However, the assumption that more fibrotic material is directly related to decreased function has not held true. Here we utilize novel measurement of extracellular matrix (ECM) and collagen architecture to relate ECM form to muscle function. We used mdx mice, a model for Duchenne muscular dystrophy that becomes fibrotic, and wildtype mice. In this model, extensor digitorum longus (EDL) muscle was significantly stiffer, but with similar total collagen, while the soleus muscle did not change stiffness, but increased collagen. The stiffness of the EDL was associated with increased collagen crosslinking as determined by collagen solubility. Measurement of ECM alignment using polarized light microscopy showed a robust relationship between stiffness and alignment for wildtype muscle that broke down in mdx muscles. Direct visualization of large collagen fibres with second harmonic generation imaging revealed their relative abundance in stiff muscles. Collagen fibre alignment was linked to stiffness across all muscles investigated and the most significant factor in a multiple linear regression-based model of muscle stiffness from ECM parameters. This work establishes novel characteristics of skeletal muscle ECM architecture and provides evidence for a mechanical function of collagen fibres in muscle. This finding suggests that anti-fibrotic strategies to enhance muscle function and excessive stiffness should target large collagen fibres and their alignment rather than total collagen.

The extracellular matrix (ECM) is a complex mixture composed of fibrillar collagens as well as additional protein and carbohydrate components. Proteoglycans (PGs) contribute to the heterogeneity of the ECM and play an important role in its structure and function. While the small leucine rich proteoglycans (SLRPs), including decorin and lumican, have been studied extensively as mediators of collagen fibrillogenesis and organization, the function of large matrix PGs in collagen matrices is less well known. In this study, we showed that different matrix PGs have distinct roles in regulating collagen behaviors. We found that versican, a large chondroitin sulfate PG, promotes collagen fibrillogenesis in a turbidity assay and upregulates cell-mediated collagen compaction and reorganization, whereas aggrecan, a structurally-similar large PG, has different and often opposing effects on collagen. Compared to versican, decorin and lumican also have distinct functions in regulating collagen behaviors. The different ways in which matrix PGs interact with collagen have important implications for understanding the role of the ECM in diseases such as fibrosis and cancer, and suggest that matrix PGs are potential therapeutic targets.

Abstract We previously demonstrated that P. aeruginosa isolates that persisted in children with cystic fibrosis (CF) despite inhaled tobramycin treatment had increased anti-Psl antibody binding in vitro compared to those successfully eradicated. We aimed to validate these findings by directly visualizing P. aeruginosa in CF sputum. This was a prospective observational study of children with CF with new-onset P. aeruginosa infection who underwent inhaled tobramycin eradication treatment. Using microbial identification passive clarity technique (MiPACT), P. aeruginosa was visualized in sputum samples obtained before treatment and classified as persistent or eradicated based on outcomes. Pre-treatment isolates were also grown as biofilms in vitro. Of 11 patients enrolled, 4 developed persistent infection and 7 eradicated infection. P. aeruginosa biovolume and the number as well as size of P. aeruginosa aggregates were greater in the sputum of those with persistent compared with eradicated infections (p

An unusual feature of the opportunistic pathogen Candida albicans is its ability to switch stochastically between two distinct, heritable cell types called white and opaque. Here, we show that only opaque cells, in response to environmental signals, massively upregulate a specific group of secreted proteases and peptide transporters, allowing exceptionally efficient use of proteins as sources of nitrogen. We identify the specific proteases [members of the secreted aspartyl protease (SAP) family] needed for opaque cells to proliferate under these conditions, and we identify four transcriptional regulators of this specialized proteolysis and uptake program. We also show that, in mixed cultures, opaque cells enable white cells to also proliferate efficiently when proteins are the sole nitrogen source. Based on these observations, we suggest that one role of white-opaque switching is to create mixed populations where the different phenotypes derived from a single genome are shared between two distinct cell types.

The fungal species Candida albicans is both a member of the human microbiome and a fungal pathogen. C. albicans undergoes several different morphological transitions, including one called white-opaque switching. Here, cells reversibly switch between two states, "white" and "opaque," and each state is heritable through many cell generations. Each cell type has a distinct cellular and colony morphology and they differ in many other properties including mating, nutritional specialization, and interactions with the innate immune system. Previous genetic screens to gain insight into white-opaque switching have focused on certain classes of genes (for example transcriptional regulators or chromatin modifying enzymes). In this paper, we examined 172 deletion mutants covering a broad range of cell functions. We identified 28 deletion mutants with at least a fivefold effect on switching frequencies; these cover a wide variety of functions ranging from membrane sensors to kinases to proteins of unknown function. In agreement with previous reports, we found that components of the pheromone signaling cascade affect white-to-opaque switching; however, our results suggest that the major effect of Cek1 on white-opaque switching occurs through the cell wall damage response pathway. Most of the genes we identified have not been previously implicated in white-opaque switching and serve as entry points to understand new aspects of this morphological transition.