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The occurrence of bacterial pathogens in the food chain has caused a severe impact on public health and welfare in both developing and developed countries. Moreover, the existence of antimicrobial-tolerant persisting morphotypes of these pathogens including both persister-cells as well as bacterial spores contributes to difficulty in elimination and in recurrent infection. Therefore, comprehensive understanding of the behavior of these persisting bacterial forms in their environmental niche and upon infection of humans is necessary. Since traditional antimicrobials fail to kill persisters and spores due to their (extremely) low metabolic activities, antimicrobial peptides (AMPs) have been intensively investigated as one of the most promising strategies against these persisting bacterial forms, showing high efficacy of inactivation. In addition, AMP-based foodborne pathogen detection and prevention of infection has made significant progress. This review focuses on recent research on common bacterial pathogens in the food chain, their persisting morphotypes, and on AMP-based solutions. Challenges in research and application of AMPs are described.

Lysosome associated membrane proteins (LAMPs) are involved in several processes, among which is fusion of lysosomes with phagosomes. For the formation of multinucleated osteoclasts, the interaction between receptor activator of nuclear kappa &beta, (RANK) and its ligand RANKL is essential. Osteoclast precursors express RANK on their membrane and RANKL is expressed by cells of the osteoblast lineage. Recently it has been suggested that the transport of RANKL to the plasma membrane is mediated by lysosomal organelles. We wondered whether LAMP-2 might play a role in transportation of RANKL to the plasma membrane of osteoblasts. To elucidate the possible function of LAMP-2 herein and in the formation of osteoclasts, we analyzed these processes in vivo and in vitro using LAMP-2-deficient mice. We found that, in the presence of macrophage colony stimulating factor (M-CSF) and RANKL, active osteoclasts were formed using bone marrow cells from calvaria and long bone mouse bone marrow. Surprisingly, an almost complete absence of osteoclast formation was found when osteoclast precursors were co-cultured with LAMP-2 deficient osteoblasts. Fluorescence-activated cell sorting FACS analysis revealed that plasma membrane-bound RANKL was strongly decreased on LAMP-2 deficient osteoblasts. These results suggest that osteoblastic LAMP-2 is required for osteoblast-induced osteoclast formation in vitro.

Transient receptor potential ankyrin 1 (TRPA1) is a membrane-bound ion channel found in neurons, where it mediates nociception and neurogenic inflammation. Recently, we have discovered that TRPA1 is also expressed in human osteoarthritic (OA) chondrocytes and downregulated by the anti-inflammatory drugs aurothiomalate and dexamethasone. We have also shown TRPA1 to mediate inflammation, pain, and cartilage degeneration in experimental osteoarthritis. In this study, we investigated the role of TRPA1 in joint inflammation, focusing on the pro-inflammatory cytokine interleukin-6 (IL-6). We utilized cartilage/chondrocytes from wild-type (WT) and TRPA1 knockout (KO) mice, along with primary chondrocytes from OA patients. The results show that TRPA1 regulates the synthesis of the OA-driving inflammatory cytokine IL-6 in chondrocytes. IL-6 was highly expressed in WT chondrocytes, and its expression, along with the expression of IL-6 family cytokines leukemia inhibitory factor (LIF) and IL-11, were significantly downregulated by TRPA1 deficiency. Furthermore, treatment with the TRPA1 antagonist significantly downregulated the expression of IL-6 in chondrocytes from WT mice and OA patients. The results suggest that TRPA1 is involved in the upregulation of IL-6 production in chondrocytes. These findings together with previous results on the expression and functions of TRPA1 in cellular and animal models point to the role of TRPA1 as a potential mediator and novel drug target in osteoarthritis. publishedVersion

Plasma membrane (PM) hyperpolarization, increased intracellular pH (pHi), and changes in intracellular calcium concentration ([Ca2+]i) are physiological events that occur during human sperm capacitation. These parameters are potential predictors of successful outcomes for men undergoing artificial reproduction techniques (ARTs), but methods currently available for their determination pose various technical challenges and limitations. Here, we developed a novel strategy employing time-lapse flow cytometry (TLFC) to determine capacitation-related membrane potential (Em) and pHi changes, and progesterone-induced [Ca2+]i increases. Our results show that TLFC is a robust method to measure absolute Em and pHi values and to qualitatively evaluate [Ca2+]i changes. To support the usefulness of our methodology, we used sperm from two types of normozoospermic donors: known paternity (subjects with self-reported paternity) and no-known paternity (subjects without self-reported paternity and no known fertility problems). We found relevant differences between them. The incidences of membrane hyperpolarization, pHi alkalinization, and increased [Ca2+]i were consistently high among known paternity samples (100%, 100%, and 86%, respectively), while they varied widely among no-known paternity samples (44%, 17%, and 45%, respectively). Our results indicate that TLFC is a powerful tool to analyze key physiological parameters of human sperm, which pending clinical validation, could potentially be employed as fertility predictors.

Streptococcus mutans is the main early colonizing cariogenic bacteria because it recognizes salivary pellicle receptors. The Antigen I/II (Ag I/II) of S. mutans is among the most important adhesins in this process, and is involved in the adhesion to the tooth surface and the bacterial co-aggregation in the early stage of biofilm formation. However, this protein has not been used as a target in a virtual strategy search for inhibitors. Based on the predicted binding affinities, drug-like properties and toxicity, molecules were selected and evaluated for their ability to reduce S. mutans adhesion. A virtual screening of 883,551 molecules was conducted, cytotoxicity analysis on fibroblast cells, S. mutans adhesion studies, scanning electron microscopy analysis for bacterial integrity and molecular dynamics simulation were also performed. We found three molecules ZINC19835187 (ZI-187), ZINC19924939 (ZI-939) and ZINC19924906 (ZI-906) without cytotoxic activity, which inhibited about 90% the adhesion of S. mutans to polystyrene microplates. Molecular dynamic simulation by 300 nanoseconds showed stability of the interaction between ZI-187 and Ag I/II (PDB: 3IPK). This work provides new molecules that targets Ag I/II and have the capacity to inhibit in vitro the S. mutans adhesion on polystyrene microplates.

Liver cirrhosis is one of the most prevalent chronic liver diseases worldwide. In addition to viral hepatitis, diseases such as steatohepatitis, autoimmune hepatitis, sclerosing cholangitis and Wilson’s disease can also lead to cirrhosis. Moreover, alcohol can cause cirrhosis on its own and exacerbate chronic liver disease of other causes. The treatment of cirrhosis can be divided into addressing the cause of cirrhosis and reversing liver fibrosis. To this date, there is still no clear consensus on the treatment of cirrhosis. Recently, there has been a lot of interest in potential treatments that modulate the gut microbiota and gut-liver axis for the treatment of cirrhosis. According to recent studies, modulation of the gut microbiome by probiotics ameliorates the progression of liver disease. The precise mechanism for relieving cirrhosis via gut microbial modulation has not been identified. This paper summarizes the role and effects of the gut microbiome in cirrhosis based on experimental and clinical studies on absorbable antibiotics, probiotics, prebiotics, and synbiotics. Moreover, it provides evidence of a relationship between the gut microbiome and liver fibrosis.

Beetle luciferases produce bioluminescence (BL) colors ranging from green to red, having been extensively used for many bioanalytical purposes, including bioimaging of pathogen infections and metastasis proliferation in living animal models and cell culture. For bioimaging purposes in mammalian tissues, red bioluminescence is preferred, due to the lower self-absorption of light at longer wavelengths by hemoglobin, myoglobin and melanin. Red bioluminescence is naturally produced only by Phrixothrix hirtus railroad worm luciferase (PxRE), and by some engineered beetle luciferases. However, Far-Red (FR) and Near-Infrared (NIR) bioluminescence is best suited for bioimaging in mammalian tissues due to its higher penetrability. Although some FR and NIR emitting luciferin analogs have been already developed, they usually emit much lower bioluminescence activity when compared to the original luciferin-luciferases. Using site-directed mutagenesis of PxRE luciferase in combination with 6&prime, modified amino-luciferin analogs, we finally selected novel FR combinations displaying BL ranging from 636&ndash, 655 nm. Among them, the combination of PxRE-R215K mutant with 6&prime, (1-pyrrolidinyl)luciferin proved to be the best combination, displaying the highest BL activity with a catalytic efficiency ~2.5 times higher than the combination with native firefly luciferin, producing the second most FR-shifted bioluminescence (650 nm), being several orders of magnitude brighter than commercial AkaLumine with firefly luciferase. Such combination also showed higher thermostability, slower BL decay time and better penetrability across bacterial cell membranes, resulting in ~3 times higher in vivo BL activity in bacterial cells than with firefly luciferin. Overall, this is the brightest FR emitting combination ever reported, and is very promising for bioimaging purposes in mammalian tissues.

C and N are the most important essential elements constituting organic compounds in plants. The shoots and roots depend on each other by exchanging C and N through the xylem and phloem transport systems. Complex mechanisms regulate C and N metabolism to optimize plant growth, agricultural crop production, and maintenance of the agroecosystem. In this paper, we cover the recent advances in understanding C and N metabolism, regulation, and transport in plants, as well as their underlying molecular mechanisms. Special emphasis is given to the mechanisms of starch metabolism in plastids and the changes in responses to environmental stress that were previously overlooked, since these changes provide an essential store of C that fuels plant metabolism and growth. We present general insights into the system biology approaches that have expanded our understanding of core biological questions related to C and N metabolism. Finally, this review synthesizes recent advances in our understanding of the trade-off concept that links C and N status to the plant’s response to microorganisms.

Dicarbonyl compounds, including methylglyoxal (MGO) and glyoxal (GO), are mainly formed as byproducts of glucose metabolism. The main glyoxalase system consists of glyoxalase I and II (Glo1 and Glo2) and is the main enzyme involved in the detoxification of dicarbonyl stress, which occurs as an accumulation of MGO or GO due to decreased activity or expression of Glo1. Dicarbonyl stress is a major cause of cellular and tissue dysfunction that causes various health issues, including diabetes, aging, and cancer. The skin is the largest organ in the body. In this review, we discuss the role of the glyoxalase system in the progression of skin aging, and more importantly, skin malignancies. We also discuss the future prospects of the glyoxalase system in other skin abnormalities such as psoriasis and vitiligo, including hyperpigmentation. Finally, in the present review, we suggest the role of glyoxalase in the progression of skin aging and glyoxalase system as a potential target for anticancer drug development for skin cancer.

Galectins are a family of animal lectins with high affinity for &beta, galactosides. Galectins are able to bind to bacteria, and a few mammalian galectins are known to kill the bound bacteria. In fish, no galectins with direct bactericidal effect have been reported. In the present study, we identified and characterized a tandem repeat galectin-8 from tongue sole Cynoglossus semilaevis (designated CsGal-8). CsGal-8 possesses conserved carbohydrate recognition domains (CRDs), as well as the conserved HXNPR and WGXEE motifs that are critical for carbohydrate binding. CsGal-8 was constitutively expressed in nine tissues of tongue sole and up-regulated in kidney, spleen, and blood by bacterial challenge. When expressed in HeLa cells, CsGal-8 protein was detected both in the cytoplasm and in the micro-vesicles secreted from the cells. Recombinant CsGal-8 (rCsGal-8) bound to lactose and other carbohydrates in a dose dependent manner. rCsGal-8 bound to a wide range of gram-positive and gram-negative bacteria and was co-localized with the bound bacteria in animal cells. Lactose, fructose, galactose, and trehalose effectively blocked the interactions between rCsGal-8 and different bacteria. Furthermore, rCsGal-8 exerted potent bactericidal activity against some gram-negative bacterial pathogens by directly damaging the membrane and structure of the pathogens. Taken together, these results indicate that CsGal-8 likely plays an important role in the immune defense against some bacterial pathogens by direct bacterial interaction and killing.

Since murine cytomegalovirus (MCMV) was first described in 1954, it has been used to model human cytomegalovirus (HCMV) diseases. MCMV is a natural pathogen of mice that is present in wild mice populations and has been associated with diseases such as myocarditis. The species-specific nature of HCMV restricts most research to cell culture-based studies or to the investigation of non-invasive clinical samples, which may not be ideal for the study of disseminated disease. Initial MCMV research used a salivary gland-propagated virus administered via different routes of inoculation into a variety of mouse strains. This revealed that the genetic background of the laboratory mice affected the severity of disease and altered the extent of subsequent pathology. The advent of genetically modified mice and viruses has allowed new aspects of disease to be modeled and the opportunistic nature of HCMV infection to be confirmed. This review describes the different ways that MCMV has been used to model HCMV diseases and explores the continuing difficulty faced by researchers attempting to model HCMV congenital cytomegalovirus disease using the mouse model.

Taro corms contain valuable bioactive molecules effective against cancer and cancer-related risk factors, such as carcinogens and biological agents, several pathophysiological conditions, including oxidative stress and inflammation, while controlling metabolic dysfunctions and boosting the immunological response. Such broad effects are achieved by the taro health-influencing compounds displaying antitumoral, antimutagenic, immunomodulatory, anti-inflammatory, antioxidant, anti-hyperglycemic, and anti-hyperlipidemic activities. Taro bioactivities are attributed to the combination of tarin, taro-4-I polysaccharide, taro polysaccharides 1 and 2 (TPS-1 and TPS-2), A-1/B-2 α-amylase inhibitors, monogalactosyldiacylglycerols (MGDGs), digalactosyldiacylglycerols (DGDGs), polyphenols, and nonphenolic antioxidants. Most of these compounds have been purified and successfully challenged in vitro and in vivo, proving their involvement in the aforementioned activities. Although these health-promoting effects have been recognized since ancient times, as well as other valuable features of taro for food profit, such as hypo-allergenicity, gluten-free, and carbohydrates with medium-glycemic index, taro crop remains underexploited. The popularization of taro intake should be considered a dietary intervention strategy to be applied to improve the overall health status of the organism and as supportive therapy to manage tumorigenesis.

Systemic Acquired Resistance (SAR) improves immunity of plant systemic tissue after local exposure to a pathogen. Guard cells that form stomatal pores on leaf surfaces recognize bacterial pathogens via pattern recognition receptors, such as Flagellin Sensitive 2 (FLS2). However, how SAR affects stomatal immunity is not known. In this study, we aim to reveal molecular mechanisms underlying the guard cell response to SAR using multi-omics of proteins, metabolites and lipids. Arabidopsis plants previously exposed to pathogenic bacteria Pseudomonas syringae pv. tomato DC3000 (Pst) exhibit an altered stomatal response compared to control plants when they are later exposed to the bacteria. Reduced stomatal apertures of SAR primed plants lead to decreased number of bacteria in leaves. Multi-omics has revealed molecular components of SAR response specific to guard cells functions, including potential roles of reactive oxygen species (ROS) and fatty acid signaling. Our results show an increase in palmitic acid and its derivative in the primed guard cells. Palmitic acid may play a role as an activator of FLS2, which initiates stomatal immune response. Improved understanding of how SAR signals affect stomatal immunity can aid biotechnology and marker-based breeding of crops for enhanced disease resistance.

Rift Valley fever virus (RVFV) is a mosquito-transmitted virus from the Bunyaviridae family that causes high rates of mortality and morbidity in humans and ruminant animals. Previous studies indicated that DEAD-box helicase 17 (DDX17) restricts RVFV replication by recognizing two primary non-coding RNAs in the S-segment of the genome: the intergenic region (IGR) and 5&prime, non-coding region (NCR). However, we lack molecular insights into the direct binding of DDX17 with RVFV non-coding RNAs and information on the unwinding of both non-coding RNAs by DDX17. Therefore, we performed an extensive biophysical analysis of the DDX17 helicase domain (DDX17135&ndash, 555) and RVFV non-coding RNAs, IGR and 5&rsquo, NCR. The homogeneity studies using analytical ultracentrifugation indicated that DDX17135&ndash, 555, IGR, and 5&rsquo, NCR are pure. Next, we performed small-angle X-ray scattering (SAXS) experiments, which suggested that DDX17 and both RNAs are homogenous as well. SAXS analysis also demonstrated that DDX17 is globular to an extent, whereas the RNAs adopt an extended conformation in solution. Subsequently, microscale thermophoresis (MST) experiments were performed to investigate the direct binding of DDX17 to the non-coding RNAs. The MST experiments demonstrated that DDX17 binds with the IGR and 5&rsquo, NCR with a dissociation constant of 5.77 &plusmn, 0.15 &micro, M and 9.85 &plusmn, 0.11 &micro, M, respectively. As DDX17135&ndash, 555 is an RNA helicase, we next determined if it could unwind IGR and NCR. We developed a helicase assay using MST and fluorescently-labeled oligos, which suggested DDX17135&ndash, 555 can unwind both RNAs. Overall, our study provides direct evidence of DDX17135&ndash, 555 interacting with and unwinding RVFV non-coding regions.

Gleditsia triacanthos is an aggressive invasive species in Eastern Europe, producing a significant number of pods that could represent an inexhaustible resource of raw material for various applications. The aim of this study was to extract cellulose from the Gleditsia triacanthos pods, characterize it by spectrophotometric and UHPLC&ndash, DAD-ESI/MS analysis, and use it to fabricate a wound dressing that is multi-functionalized with phenolic compounds extracted from the leaves of the same species. The obtained cellulose microfibers (CM) were functionalized, lyophilized, and characterized by ATR-FTIR and SEM. The water absorption and retention capacity as well as the controlled release of phenolic compounds with antioxidant properties evaluated in temporal dynamics were also determined. The antimicrobial activity against reference and clinical multi-drug-resistant Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii, Enterobacter cloacae, Candida albicans, and Candida parapsilosis strains occurred immediately after the contact with the tested materials and was maintained for 24 h for all tested microbial strains. In conclusion, the multi-functionalized cellulose microfibers (MFCM) obtained from the reproductive organs of an invasive species can represent a promising alternative for the development of functional wound dressings with antioxidant and antimicrobial activity, as well as being a scalable example for designing cost-effective, circular bio-economy approaches to combat the accelerated spread of invasive species.

International audience; B-cell non-Hodgkin's lymphoma (NHL) risk associations had been mainly attributed to family history of the disease, inflammation, and immune components including human leukocyte antigen (HLA) genetic variations. Nevertheless, a broad range of genome-wide association studies (GWAS) have shed light into the identification of several genetic variants presumptively associated with B-cell NHL etiologies, survival or shared genetic risk with other diseases. The present review aims to overview HLA structure and diversity and summarize the evidence of genetic variations, by GWAS, on five NHL subtypes (diffuse large B-cell lymphoma DLBCL, follicular lymphoma FL, chronic lymphocytic leukemia CLL, marginal zone lymphoma MZL, and primary central nervous system lymphoma PCNSL). Evidence indicates that the HLA zygosity status in B-cell NHL might promote immune escape and that genome-wide significance variants can give biological insight but also potential therapeutic markers such as WEE1 in DLBCL. However, additional studies are needed, especially for non-DLBCL, to replicate the associations found to date.

The iron-sulfur subunit (SDH2) of succinate dehydrogenase plays a key role in electron transport in plant mitochondria. However, it is yet unknown whether SDH2 genes are involved in leaf senescence and yield formation. In this study, we isolated a late premature senescence mutant, lps1, in rice (Oryza sativa). The mutant leaves exhibited brown spots at late tillering stage and wilted at the late grain-filling stage and mature stage. In its premature senescence leaves, photosynthetic pigment contents and net photosynthetic rate were reduced, chloroplasts and mitochondria were degraded. Meanwhile, lps1 displayed small panicles, low seed-setting rate and dramatically reduced grain yield. Gene cloning and complementation analysis suggested that the causal gene for the mutant phenotype was OsSDH2-1 (LOC_Os08g02640), in which single nucleotide mutation resulted in an amino acid substitution in the encoded protein. OsSDH2-1 gene was expressed in all organs tested, with higher expression in leaves, root tips, ovary and anthers. OsSDH2-1 protein was targeted to mitochondria. Furthermore, reactive oxygen species (ROS), mainly H2O2, was excessively accumulated in leaves and young panicles of lps1, which could cause premature leaf senescence and affect panicle development and pollen function. Taken together, OsSDH2-1 plays a crucial role in leaf senescence and yield formation in rice.

The immune privilege of the testes is necessary to prevent immune attacks to gamete-specific antigens and paternal major histocompatibility complex (MHC) antigens, allowing for normal spermatogenesis. However, infection and inflammation of the male genital tract can break the immune tolerance and represent a significant cause of male infertility. Different T cell subsets have been identified in mammalian testes, which may be involved in the maintenance of immune tolerance and pathogenic immune responses in testicular infection and inflammation. We reviewed the evidence in the published literature on different T subtypes (regulatory T cells, helper T cells, cytotoxic T cells, γδ T cells, and natural killer T cells) in human and animal testes that support their regulatory roles in infertility and the orchitis pathology. While many in vitro studies have indicated the regulation potential of functional T cell subsets and their possible interaction with Sertoli cells, Leydig cells, and spermatogenesis, both under physiological and pathological processes, there have been no in situ studies to date. Nevertheless, the normal distribution and function of T cell subsets are essential for the immune privilege of the testes and intact spermatogenesis, and T cell-mediated immune response drives testicular inflammation. The distinct function of different T cell subsets in testicular homeostasis and the orchitis pathology suggests a considerable potential of targeting specific T cell subsets for therapies targeting chronic orchitis and immune infertility.

We thank Toffa (Language Advisory Service, University of Barcelona, Barcelona, Spain) for revising the English text, Microbiota is defined as the collection of microorganisms within the gastrointestinal ecosystem. These microbes are strongly implicated in the stimulation of immune responses. An unbalanced microbiota, termed dysbiosis, is related to the development of several liver diseases. The bidirectional relationship between the gut, its microbiota and the liver is referred to as the gut–liver axis. The translocation of bacterial products from the intestine to the liver induces inflammation in different cell types such as Kupffer cells, and a fibrotic response in hepatic stellate cells, resulting in deleterious effects on hepatocytes. Moreover, ischemia-reperfusion injury, a consequence of liver surgery, alters the microbiota profile, affecting inflammation, the immune response and even liver regeneration. Microbiota also seems to play an important role in post-operative outcomes (i.e., liver transplantation or liver resection). Nonetheless, studies to determine changes in the gut microbial populations produced during and after surgery, and affecting liver function and regeneration are scarce. In the present review we analyze and discuss the preclinical and clinical studies reported in the literature focused on the evaluation of alterations in microbiota and its products as well as their effects on post-operative outcomes in hepatic surgery., Ministerio de Ciencia, Innovacion y Universidades (MCIU) Madrid, Spain RTI2018-095114-B-I00, European Union (Fondos Feder, "Una manera de hacer Europa"), CERCA Program/Generalitat de Catalunya, Secretaria d'Universitats i Recerca Barcelona, Spain 2017SGR-551, COST action Programs CA17103 CA17126

Pulmonary fibrosis is one of the important causes of morbidity and mortality in fibroproliferative disorders such as systemic sclerosis (SSc) and idiopathic pulmonary fibrosis (IPF). Insulin-like growth factor binding protein-5 (IGFBP-5) is a conserved member of the IGFBP family of proteins that is overexpressed in SSc and IPF lung tissues. In this study, we investigated the functional role of IGFBP-5 in the development of fibrosis in vivo using a transgenic model. We generated transgenic mice ubiquitously expressing human IGFBP-5 using CRISPR/Cas9 knock-in. Our data show that the heterozygous and homozygous mice are viable and express human IGFBP-5 (hIGFBP-5). Transgenic mice had increased expression of extracellular matrix (ECM) genes, especially Col3a1, Fn, and Lox in lung and skin tissues of mice expressing higher transgene levels. Histologic analysis of the skin tissues showed increased dermal thickness, and the lung histology showed subtle changes in the heterozygous and homozygous mice as compared with the wild-type mice. These changes were more pronounced in animals expressing higher levels of hIGFBP-5. Bleomycin increased ECM gene expression in wild-type mice and accentuated an increase in ECM gene expression in transgenic mice, suggesting that transgene expression exacerbated bleomycin-induced pulmonary fibrosis. Primary lung fibroblasts cultured from lung tissues of homozygous transgenic mice showed significant increases in ECM gene expression and protein levels, further supporting the observation that IGFBP-5 resulted in a fibrotic phenotype in fibroblasts. In summary, transgenic mice expressing human IGFBP-5 could serve as a useful animal model for examining the function of IGFBP-5 in vivo.

Hospital-acquired infections (HAIs) are a growing concern around the world. They contribute to increasing mortality and morbidity rates and are an economic threat. All hospital patients have the potential to contract an HAI, but those with weakened or inferior immune systems are at highest risk. Most hospital patients will contract at least one HAI, but many will contract multiple ones. Bacteria are the most common cause of HAIs and contribute to 80–90% of all HAIs, with Staphylococcus aureus, Clostridium difficile, Escherichia coli, Acinetobacter baumannii, Pseudomonas aeruginosa and Klebsiella pneumoniae accounting for the majority. Each of these bacteria are highly resistant to antibiotics and can produce a protective film, known as a biofilm, to further prevent their eradication. It has been shown that by detecting and eradicating bacteria in the environment, infection rates can be reduced. The current methods for detecting bacteria are time consuming, non-specific, and prone to false negatives or false positives. Aptamer-based biosensors have demonstrated specific, time-efficient and simple detection, highlighting the likelihood that they could be used in a similar way to detect HAI-causing bacteria.

β-Endorphins are peptides that exert a wide variety of effects throughout the body. Produced through the cleavage pro-opiomelanocortin (POMC), β-endorphins are the primarily agonist of mu opioid receptors, which can be found throughout the body, brain, and cells of the immune system that regulate a diverse set of systems. As an agonist of the body’s opioid receptors, β-endorphins are most noted for their potent analgesic effects, but they also have their involvement in reward-centric and homeostasis-restoring behaviors, among other effects. These effects have implicated the peptide in psychiatric and neurodegenerative disorders, making it a research target of interest. This review briefly summarizes the basics of endorphin function, goes over the behaviors and regulatory pathways it governs, and examines the variability of β-endorphin levels observed between normal and disease/disorder affected individuals.

Pseudomonas putida&rsquo, s versatility and metabolic flexibility make it an ideal biotechnological platform for producing valuable chemicals, such as medium-chain-length polyhydroxyalkanoates (mcl-PHAs), which are considered the next generation bioplastics. This bacterium responds to environmental stimuli by rearranging its metabolism to improve its fitness and increase its chances of survival in harsh environments. Mcl-PHAs play an important role in central metabolism, serving as a reservoir of carbon and energy. Due to the complexity of mcl-PHAs&rsquo, metabolism, the manner in which P. putida changes its transcriptome to favor mcl-PHA synthesis in response to environmental stimuli remains unclear. Therefore, our objective was to investigate how the P. putida KT2440 wild type and mutants adjust their transcriptomes to synthesize mcl-PHAs in response to nitrogen limitation when supplied with sodium gluconate as an external carbon source. We found that, under nitrogen limitation, mcl-PHA accumulation is significantly lower in the mutant deficient in the stringent response than in the wild type or the rpoN mutant. Transcriptome analysis revealed that, under N-limiting conditions, 24 genes were downregulated and 21 were upregulated that were common to all three strains. Additionally, potential regulators of these genes were identified: the global anaerobic regulator (Anr, consisting of FnrA, Fnrb, and FnrC), NorR, NasT, the sigma54-dependent transcriptional regulator, and the dual component NtrB/NtrC regulator all appear to play important roles in transcriptome rearrangement under N-limiting conditions. The role of these regulators in mcl-PHA synthesis is discussed.

The origin of chlorophyll b deficiency is a mutation (ch1) in chlorophyllide a oxygenase (CAO), the enzyme responsible for Chl b synthesis. Regulation of Chl b synthesis is essential for understanding the mechanism of plant acclimation to various conditions. Therefore, the main aim of this study was to find the strategy in plants for compensation of low chlorophyll content by characterizing and comparing the performance and spectral properties of the photosynthetic apparatus related to the lipid and protein composition in four selected Arabidopsis ch1 mutants and two Arabidopsis ecotypes. Mutation in different loci of the CAO gene, viz., NW41, ch1.1, ch1.2 and ch1.3, manifested itself in a distinct chlorina phenotype, pigment and photosynthetic protein composition. Changes in the CAO mRNA levels and chlorophyllide a (Chlide a) content in ecotypes and ch1 mutants indicated their significant role in the adjustment mechanism of the photosynthetic apparatus to low-light conditions. Exposure of mutants with a lower chlorophyll b content to short-term (1LL) and long-term low-light stress (10LL) enabled showing a shift in the structure of the PSI and PSII complexes via spectral analysis and the thylakoid composition studies. We demonstrated that both ecotypes, Col-1 and Ler-0, reacted to high-light (HL) conditions in a way remarkably resembling the response of ch1 mutants to normal (NL) conditions. We also presented possible ways of regulating the conversion of chlorophyll a to b depending on the type of light stress conditions.

Dendroctonus-bark beetles are natural agents contributing to vital processes in coniferous forests, such as regeneration, succession, and material recycling, as they colonize and kill damaged, stressed, or old pine trees. These beetles spend most of their life cycle under stem and roots bark where they breed, develop, and feed on phloem. This tissue is rich in essential nutrients and complex molecules such as starch, cellulose, hemicellulose, and lignin, which apparently are not available for these beetles. We evaluated the digestive capacity of Dendroctonusrhizophagus to hydrolyze starch. Our aim was to identify &alpha, amylases and characterize them both molecularly and biochemically. The findings showed that D. rhizophagus has an &alpha, amylase gene (AmyDr) with a single isoform, and ORF of 1452 bp encoding a 483-amino acid protein (53.15 kDa) with a predicted signal peptide of 16 amino acids. AmyDr has a mutation in the chlorine-binding site, present in other phytophagous insects and in a marine bacterium. Docking analysis showed that AmyDr presents a higher binding affinity to amylopectin compared to amylose, and an affinity binding equally stable to calcium, chlorine, and nitrate ions. AmyDr native protein showed amylolytic activity in the head-pronotum and gut, and its recombinant protein, a polypeptide of ~53 kDa, showed conformational stability, and its activity is maintained both in the presence and absence of chlorine and nitrate ions. The AmyDr gene showed a differential expression significantly higher in the gut than the head-pronotum, indicating that starch hydrolysis occurs mainly in the midgut. An overview of the AmyDr gene expression suggests that the amylolytic activity is regulated through the developmental stages of this bark beetle and associated with starch availability in the host tree.

Rice (Oryza sativa L.) is one of the major crops in the world and significant increase in grain yield is constant demand for breeders to meet the needs of a rapidly growing population. The size of grains is one of major components determining rice yield and a vital trait for domestication and breeding. To increase the grain size in rice, OsSPL16/qGW8 was mutagenized through CRISPR/Cas9, and proteomic analysis was performed to reveal variations triggered by mutations. More specifically, mutants were generated with two separate guide RNAs targeting recognition sites on opposite strands and genomic insertions and deletions were characterized. Mutations followed Mendelian inheritance and homozygous and heterozygous mutants lacking any T-DNA and off-target effects were screened. The mutant lines showed a significant increase in grain yield without any change in other agronomic traits in T0, T1, and T2 generations. Proteomic screening found a total of 44 differentially expressed proteins (DEPs), out of which 33 and 11 were up and downregulated, respectively. Most of the DEPs related to pyruvate kinase, pyruvate dehydrogenase, and cell division and proliferation were upregulated in the mutant plants. Pathway analysis revealed that DEPs were enriched in the biosynthesis of secondary metabolites, pyruvate metabolism, glycolysis/gluconeogenesis, carbon metabolism, ubiquinone and other terpenoid-quinone biosynthesis, and citrate cycle. Gene Ontology (GO) analysis presented that most of the DEPs were involved in the pyruvate metabolic process and pyruvate dehydrogenase complex. Proteins related to pyruvate dehydrogenase E1 component subunit alpha-1 displayed higher interaction in the protein-protein interaction (PPI) network. Thus, the overall results revealed that CRISPR/Cas9-guided OsSPL16 mutations have the potential to boost the grain yield of rice. Additionally, global proteome analysis has broad applications for discovering molecular components and dynamic regulation underlying the targeted gene mutations.

The gastrointestinal tract (GIT) plays a key role in regulating nutrient metabolism and appetite responses. This study aimed to identify changes in the GIT that are important in the development of diet related obesity and diabetes. GIT samples were obtained from C57BL/6J male mice chronically fed a control diet or a high sucrose diet (HSD) and analysed for changes in gene, protein and metabolite levels. In HSD mice, GIT expression levels of fat oxidation genes were reduced, and increased de novo lipogenesis was evident in ileum. Gene expression levels of the putative sugar sensor, slc5a4a and slc5a4b, and fat sensor, cd36, were downregulated in the small intestines of HSD mice. In HSD mice, there was also evidence of bacterial overgrowth and a lipopolysaccharide activated inflammatory pathway involving inducible nitric oxide synthase (iNOS). In Caco-2 cells, sucrose significantly increased the expression levels of the nos2, iNOS and nitric oxide (NO) gas levels. In conclusion, sucrose fed induced obesity/diabetes is associated with changes in GI macronutrient sensing, appetite regulation and nutrient metabolism and intestinal microflora. These may be important drivers, and thus therapeutic targets, of diet-related metabolic disease.

There is increasing recognition that environmental nano-biological interactions in model species, and the resulting effects on progeny, are of paramount importance for nanomaterial (NM) risk assessment. In this work, Daphnia magna F0 mothers were exposed to a range of silver and titanium dioxide NMs. The key biological life history traits (survival, growth and reproduction) of the F1 intergenerations, at the first (F1B1), third (F1B3) and fifth (F1B5) broods, were investigated. Furthermore, the F1 germlines of each of the three broods were investigated over 3 more generations (up to 25 days each) in continuous or removed-from NM exposure, to identify how the length of maternal exposure affects the resulting clonal broods. Our results show how daphnids respond to NM-induced stress, and how the maternal effects show trade-offs between growth, reproduction and survivorship. The F1B1 (and following germline) had the shortest F0 maternal exposure times to the NMs, and thus were the most sensitive showing reduced size and reproductive output. The F1B3 generation had a sub-chronic maternal exposure, whereas the F1B5 generation suffered chronic maternal exposure where (in most cases) the most compensatory adaptive effects were displayed in response to the prolonged NM exposure, including enhanced neonate output and reduced gene expression. Transgenerational responses of multiple germlines showed a direct link with maternal exposure time to &lsquo, sub-lethal&rsquo, effect concentrations of NMs (identified from standard OECDs acute toxicity tests which chronically presented as lethal) including increased survival and production of males in the F1B3 and G1B5 germlines. This information may help to fine-tune environmental risk assessments of NMs and prediction of their impacts on environmental ecology.

Fetal undernutrition programs cardiometabolic diseases, with higher susceptibility in males. The mechanisms implicated are not fully understood and may be related to sex differences in placental adaptation. To evaluate this hypothesis, we investigated placental oxidative balance, vascularization, glucocorticoid barrier, and fetal growth in rats exposed to 50% global nutrient restriction from gestation day 11 (MUN, n = 8) and controls (n = 8). At gestation day 20 (G20), we analyzed maternal, placental, and fetal weights, oxidative damage, antioxidants, corticosterone, and PlGF (placental growth factor, spectrophotometry), and VEGF (vascular endothelial growth factor), 11&beta, HSD2, p22phox, XO, SOD1, SOD2, SOD3, catalase, and UCP2 expression (Western blot). Compared with controls, MUN dams exhibited lower weight and plasma proteins and higher corticosterone and catalase without oxidative damage. Control male fetuses were larger than female fetuses. MUN males had higher plasma corticosterone and were smaller than control males, but had similar weight than MUN females. MUN male placenta showed higher XO and lower 11&beta, HSD2, VEGF, SOD2, catalase, UCP2, and feto-placental ratio than controls. MUN females had similar feto-placental ratio and plasma corticosterone than controls. Female placenta expressed lower XO, 11&beta, HSD2, and SOD3, similar VEGF, SOD1, SOD2, and UCP2, and higher catalase than controls, being 11&beta, HSD2 and VEGF higher compared to MUN males. Male placenta has worse adaptation to undernutrition with lower efficiency, associated with oxidative disbalance and reduced vascularization and glucocorticoid barrier. Glucocorticoids and low nutrients may both contribute to programming in MUN males.

In order to reduce the need for donor corneas, understanding of corneal wound healing and development of an entirely tissue-engineered human cornea (hTECs) is of prime importance. In this study, we exploited the hTEC to determine how deep wound healing affects the transcriptional pattern of corneal epithelial cells through microarray analyses. We demonstrated that the gene encoding clusterin (CLU) has its expression dramatically repressed during closure of hTEC wounds. Western blot analyses confirmed a strong reduction in the expression of the clusterin isoforms after corneal damage and suggest that repression of CLU gene expression might be a prerequisite to hTEC wound closure. Transfection with segments from the human CLU gene promoter revealed the presence of three regulatory regions: a basal promoter and two more distal negative regulatory regions. The basal promoter bears DNA binding sites for very potent transcription factors (TFs): Activator Protein-1 (AP-1) and Specificity protein-1 and 3 (Sp1/Sp3). By exploiting electrophoretic mobility shift assays (EMSA), we demonstrated that AP-1 and Sp1/Sp3 have their DNA binding site overlapping with one another in the basal promoter of the CLU gene in hCECs. Interestingly, expression of both these TFs is reduced (at the protein level) during hTEC wound healing, thereby contributing to the extinction of CLU gene expression during that process. The results of this study contribute to a better understanding of the molecular mechanisms accounting for the repression of CLU gene expression during corneal wound healing.

DNA molecules can adopt a variety of alternative structures. Among these structures are G-quadruplex DNA structures (G4s), which support cellular function by affecting transcription, translation, and telomere maintenance. These structures can also induce genome instability by stalling replication, increasing DNA damage, and recombination events. G-quadruplex-driven genome instability is connected to tumorigenesis and other genetic disorders. In recent years, the connection between genome stability, DNA repair and G4 formation was further underlined by the identification of multiple DNA repair proteins and ligands which bind and stabilize said G4 structures to block specific DNA repair pathways. The relevance of G4s for different DNA repair pathways is complex and depends on the repair pathway itself. G4 structures can induce DNA damage and block efficient DNA repair, but they can also support the activity and function of certain repair pathways. In this review, we highlight the roles and consequences of G4 DNA structures for DNA repair initiation, processing, and the efficiency of various DNA repair pathways.

Dystrophin is a 427 kDa protein that stabilizes muscle cell membranes through interactions with the cytoskeleton and various membrane-associated proteins. Loss of dystrophin as in Duchenne muscular dystrophy (DMD) causes progressive skeletal muscle weakness and cardiac dysfunction. Multiple promoters along the dystrophin gene (DMD) give rise to a number of shorter isoforms. Of interest is Dp71, a 71 kDa isoform implicated in DMD pathology by various animal and patient studies. Strong evidence supporting such a role for Dp71, however, is lacking. Here, we use del52;WT mice to understand how Dp71 overexpression affects skeletal and cardiac muscle phenotypes. Apart from the mouse Dmd gene, del52;WT mice are heterozygous for a full-length, exon 52-deleted human DMD transgene expected to only permit Dp71 expression in muscle. Thus, del52;WT mice overexpress Dp71 through both the human and murine dystrophin genes. We observed elevated Dp71 protein in del52;WT mice, significantly higher than wild-type in the heart but not the tibialis anterior. Moreover, del52;WT mice had generally normal skeletal muscle but impaired cardiac function, exhibiting significant systolic dysfunction as early as 3 months. No histological abnormalities were found in the tibialis anterior and heart. Our results suggest that Dp71 overexpression may have more detrimental effects on the heart than on skeletal muscles, providing insight into the role of Dp71 in DMD pathogenesis.

Fungal pathogens cause an array of diseases by targeting both immunocompromised and immunocompetent hosts. Fungi overcome our current arsenal of antifungals through the emergence and evolution of resistance. In particular, the human fungal pathogen, Cryptococcus neoformans is found ubiquitously within the environment and causes severe disease in immunocompromised individuals around the globe with limited treatment options available. To uncover fundamental knowledge about this fungal pathogen, as well as investigate new detection and treatment strategies, mass spectrometry-based proteomics provides a plethora of tools and applications, as well as bioinformatics platforms. In this review, we highlight proteomics approaches within the laboratory to investigate changes in the cellular proteome, secretome, and extracellular vesicles. We also explore regulation by post-translational modifications and the impact of protein–protein interactions. Further, we present the development and comprehensive assessment of murine models of cryptococcal infection, which provide valuable tools to define the dynamic relationship between the host and pathogen during disease. Finally, we explore recent quantitative proteomics studies that begin to extrapolate the findings from the bench to the clinic for improved methods of fungal detection and monitoring. Such studies support a framework for personalized medical approaches to eradicate diseases caused by C. neoformans.

Tissue homeostasis via the elimination of aberrant cells is fundamental for organism survival. Cell competition is a key homeostatic mechanism, contributing to the recognition and elimination of aberrant cells, preventing their malignant progression and the development of tumors. Here, using Drosophila as a model organism, we have defined a role for protein tyrosine phosphatase 61F (PTP61F) (orthologue of mammalian PTP1B and TCPTP) in the initiation and progression of epithelial cancers. We demonstrate that a Ptp61F null mutation confers cells with a competitive advantage relative to neighbouring wild-type cells, while elevating PTP61F levels has the opposite effect. Furthermore, we show that knockdown of Ptp61F affects the survival of clones with impaired cell polarity, and that this occurs through regulation of the JAK–STAT signalling pathway. Importantly, PTP61F plays a robust non-cell-autonomous role in influencing the elimination of adjacent polarity-impaired mutant cells. Moreover, in a neoplastic RAS-driven polarity-impaired tumor model, we show that PTP61F levels determine the aggressiveness of tumors, with Ptp61F knockdown or overexpression, respectively, increasing or reducing tumor size. These effects correlate with the regulation of the RAS–MAPK and JAK–STAT signalling by PTP61F. Thus, PTP61F acts as a tumor suppressor that can function in an autonomous and non-cell-autonomous manner to ensure cellular fitness and attenuate tumorigenesis.

International audience; Visual deficit is one of the complications of Huntington disease (HD), a fatal neurological disorder caused by CAG trinucleotide expansions in the Huntingtin gene, leading to the production of mutant Huntingtin (mHTT) protein. Transgenic HD R6/1 mice expressing human HTT exon1 with 115 CAG repeats recapitulate major features of the human pathology and exhibit a degeneration of the retina. Our aim was to gain insight into the ultrastructure of the pathological HD R6/1 retina by electron microscopy (EM). We show that the HD R6/1 retina is enriched with unusual organelles myelinosomes, produced by retinal neurons and glia. Myelinosomes are present in all nuclear and plexiform layers, in the synaptic terminals of photoreceptors, in the processes of retinal neurons and glial cells, and in the subretinal space. In vitro study shows that myelinosomes secreted by human retinal glial Müller MIO-M1 cells transfected with EGFP-mHTT-exon1 carry EGFP-mHTT-exon1 protein, as revealed by immuno-EM and Western-blotting. Myelinosomes loaded with mHTT-exon1 are incorporated by naive neuronal/neuroblastoma SH-SY5Y cells. This results in the emergence of mHTT-exon1 in recipient cells. This process is blocked by membrane fusion inhibitor MDL 28170. Conclusion. Incorporation of myelinosomes carrying mHTT-exon1 in recipient cells may contribute to HD spreading in the retina. Exploring ocular fluids for myelinosome presence could bring an additional biomarker for HD diagnostics. © 2021 by the authors. Licensee MDPI, Basel, Switzerland.

Recessive dystrophic epidermolysis bullosa (RDEB) is a rare autosomal inherited skin disorder caused by mutations in the COL7A1 gene that encodes type VII collagen (C7). The development of an efficient gene replacement strategy for RDEB is mainly hindered by the lack of vectors able to encapsulate and transfect the large cDNA size of this gene. To address this problem, our group has opted to use polymeric-based non-viral delivery systems and minicircle DNA. With this approach, safety is improved by avoiding the usage of viruses, the absence of bacterial backbone, and the replacement of the control viral cytomegalovirus (CMV) promoter of the gene with human promoters. All the promoters showed impressive C7 expression in RDEB skin cells, with eukaryotic translation elongation factor 1 α (EF1α) promoter producing higher C7 expression levels than CMV following minicircle induction, and COL7A1 tissue-specific promoter (C7P) generating C7 levels similar to normal human epidermal keratinocytes. The improved system developed here has a high potential for use as a non-viral topical treatment to restore C7 in RDEB patients efficiently and safely, and to be adapted to other genetic conditions.

The early-life microbiome (ELM) interacts with the psychosocial environment, in particular during early-life adversity (ELA), defining life-long health trajectories. The ELM also plays a significant role in the maturation of the immune system. We hypothesised that, in this context, the resilience of the oral microbiomes, despite being composed of diverse and distinct communities, allows them to retain an imprint of the early environment. Using 16S amplicon sequencing on the EpiPath cohort, we demonstrate that ELA leaves an imprint on both the salivary and buccal oral microbiome 24 years after exposure to adversity. Furthermore, the changes in both communities were associated with increased activation, maturation, and senescence of both innate and adaptive immune cells, although the interaction was partly dependent on prior herpesviridae exposure and current smoking. Our data suggest the presence of multiple links between ELA, Immunosenescence, and cytotoxicity that occur through long-term changes in the microbiome.

The human small intestine can be involved in the first-pass metabolism of drugs. Under this condition, members of the CYP450 superfamily are expected to contribute to drug presystemic biotransformation. The aim of this study was to quantify protein expression levels of 16 major CYP450 isoforms in tissue obtained from nine human organ donors in seven subsections of the small intestine, i.e., duodenum (one section, N = 7 tissue samples), jejunum (three subsections (proximal, mid and distal), N = 9 tissue samples) and ileum (three subsections, (proximal, mid and distal), N = 9 tissue samples), using liquid chromatography tandem mass spectrometry (LC-MS/MS) based targeted proteomics. CYP450 absolute protein expression levels were compared to mRNA levels and enzyme activities by using established probe drugs. Proteins corresponding to seven of sixteen potential CYP450 isoforms were detected and quantified in various sections of the small intestine: CYP2C9, CYP2C19, CYP2D6, CYP2J2, CYP3A4, CYP3A5 and CYP4F2. Wide inter-subject variability was observed, especially for CYP2D6. CYP2C9 (p = 0.004) and CYP2C19 (p = 0.005) expression levels decreased along the small intestine. From the duodenum to the ileum, CYP2J2 (p = 0.001) increased, and a trend was observed for CYP3A5 (p = 0.13). CYP3A4 expression was higher in the jejunum than in the ileum (p = 0.03), while CYP4F2 expression was lower in the duodenum compared to the jejunum and the ileum (p = 0.005). CYP450 protein levels were better correlated with specific isoform activities than with mRNA levels. This study provides new data on absolute CYP450 quantification in human small intestine that could improve physiologically based pharmacokinetic models. These data could better inform drug absorption profiles while considering the regional expression of CYP450 isoforms.

The opportunistic pathogen Pseudomonas aeruginosa is a significant cause of infection in immunocompromised individuals, cystic fibrosis patients, and burn victims. To benefit its survival, the bacterium adapt to either a motile or sessile lifestyle when infecting the host. The motile bacterium has an often activated type III secretion system (T3SS), which is virulent to the host, whereas the sessile bacterium harbors an active T6SS and lives in biofilms. Regulatory pathways involving Gac-Rsm or secondary messengers such as c-di-GMP determine which lifestyle is favorable for P. aeruginosa. Here, we introduce the RNA binding protein RtcB as a modulator of the switch between motile and sessile bacterial lifestyles. Using the wild-type P. aeruginosa PAO1, and a retS mutant PAO1(∆retS) in which T3SS is repressed and T6SS active, we show that deleting rtcB led to simultaneous expression of T3SS and T6SS in both PAO1(∆rtcB) and PAO1(∆rtcB∆retS). The deletion of rtcB also increased biofilm formation in PAO1(∆rtcB) and restored the motility of PAO1(∆rtcB∆retS). RNA-sequencing data suggested RtcB as a global modulator affecting multiple virulence factors, including bacterial secretion systems. Competitive killing and infection assays showed that the three T6SS systems (H1, H2, and H3) in PAO1(∆rtcB) were activated into a functional syringe, and could compete with Escherichia coli and effectively infect lettuce. Western blotting and RT-PCR results showed that RtcB probably exerted its function through RsmA in PAO1(∆rtcB∆retS). Quantification of c-di-GMP showed an elevated intracellular levels in PAO1(∆rtcB), which likely drove the switch between T6SS and T3SS, and contributed to the altered phenotypes and characteristics observed. Our data demonstrate a pivotal role of RtcB in the virulence of P. aeruginosa by controlling multiple virulence determinants, such as biofilm formation, motility, pyocyanin production, T3SS, and T6SS secretion systems towards eukaryotic and prokaryotic cells. These findings suggest RtcB as a potential target for controlling P. aeruginosa colonization, establishment, and pathogenicity.

International journal of molecular sciences 22(21), 11779 (2021). doi:10.3390/ijms222111779 special issue: "Topical Collection "Feature Papers in Molecular Pharmacology" / Editor: Prof. Dr. Peter Illes, Collection Editor", Published by Molecular Diversity Preservation International, Basel

International audience; Microgravity, one of the conditions faced by astronauts during spaceflights, triggers brain adaptive responses that could have noxious consequences on behaviors. Although monoaminergic systems, which include noradrenaline (NA), dopamine (DA), and serotonin (5-HT), are widespread neuromodulatory systems involved in adaptive behaviors, the influence of microgravity on these systems is poorly documented. Using a model of simulated microgravity (SMG) during a short period in Long Evans male rats, we studied the distribution of monoamines in thirty brain regions belonging to vegetative, mood, motor, and cognitive networks. SMG modified NA and/or DA tissue contents along some brain regions belonging to the vestibular/motor systems (inferior olive, red nucleus, cerebellum, somatosensorily cortex, substantia nigra, and shell of the nucleus accumbens). DA and 5-HT contents were reduced in the prelimbic cortex, the only brain area exhibiting changes for 5-HT content. However, the number of correlations of one index of the 5-HT metabolism (ratio of metabolite and 5-HT) alone or in interaction with the DA metabolism was dramatically increased between brain regions. It is suggested that SMG, by mobilizing vestibular/motor systems, promotes in these systems early, restricted changes of NA and DA functions that are associated with a high reorganization of monoaminergic systems, notably 5-HT.

Epithelial-to-mesenchymal transition (EMT) recapitulates metastasis and can be induced in vitro through transforming growth factor (TGF)-β signaling. A role for MMP activity in glioblastoma multiforme has been ascribed to EMT, but the molecular crosstalk between TGF-β signaling and membrane type 1 MMP (MT1-MMP) remains poorly understood. Here, the expression of common EMT biomarkers, induced through TGF-β and the MT1-MMP inducer concanavalin A (ConA), was explored using RNA-seq analysis and differential gene arrays in human U87 glioblastoma cells. TGF-β triggered SNAIL and fibronectin expressions in 2D-adherent and 3D-spheroid U87 glioblastoma cell models. Those inductions were antagonized by the TGF-β receptor kinase inhibitor galunisertib, the JAK/STAT inhibitors AG490 and tofacitinib, and by the diet-derived epigallocatechin gallate (EGCG). Transient gene silencing of MT1-MMP prevented the induction of SNAIL by ConA and abrogated TGF-β-induced cell chemotaxis. Moreover, ConA induced STAT3 and Src phosphorylation, suggesting these pathways to be involved in the MT1-MMP-mediated signaling axis that led to SNAIL induction. Our findings highlight a new signaling axis linking MT1-MMP to TGF-β-mediated EMT-like induction in glioblastoma cells, the process of which can be prevented by the diet-derived EGCG.

The goal of this study was to explore the specific signaling pathways related to inflammation in two experimental mouse dry eye (EDE) models. Female C57BL/6 mice housed for 10 days in a controlled desiccative environment were either treated with scopolamine (EDE-1, n = 18) or subjected to extraorbital lacrimal gland excision bilaterally (EDE-2, n = 10). Non-induced mice (n = 20) served as healthy controls. A corneal fluorescein staining (CFS) scoring was used at baseline through to day (D) 10 to evaluate epitheliopathy. At D10, corneas and conjunctivas were collected for multiplexed transcriptomic analysis with the NanoString® mouse inflammatory CodeSet. Both EDE-1 and EDE-2 mice presented a change in corneal integrity, with a significant increase in CFS scores at D10. More gene transcripts were identified in EDE-2 compared with EDE-1 (116 vs. 96, respectively), and only a few were common to both models, 13 for the cornea and 6 for the conjunctiva. The gene functional annotation analysis revealed that the same inflammatory pathways were involved in both models. Comparative profiling of gene expression in the two EDE models leads to the identification of various targets and signaling pathways, which can be extrapolated to and confirmed in human disease.

In recent decades, neurogenesis in the adult brain has been well demonstrated in anumber of animal species, including humans. Interestingly, work with rodents has shown that adultneurogenesis in the dentate gyrus (DG) of the hippocampus is vital for some cognitive aspects, asincreasing neurogenesis improves memory, while its disruption triggers the opposite effect. Adultneurogenesis declines with age and has been suggested to play a role in impaired progressive learningand memory loss seen in Alzheimer’s disease (AD). Therefore, therapeutic strategies designed toboost adult hippocampal neurogenesis may be beneficial for the treatment of AD. The precursor formsof neurotrophins, such as pro-NGF, display remarkable increase during AD in the hippocampusand entorhinal cortex. In contrast to mature NGF, pro-NGF exerts adverse functions in survival,proliferation, and differentiation. Hence, we hypothesized that pro-NGF and its p75 neurotrophinreceptor (p75NTR) contribute to disrupting adult hippocampal neurogenesis during AD. To test thishypothesis, in this study, we took advantage of the availability of mouse models of AD (APP/PS1),which display memory impairment, and AD human samples to address the role of pro-NGF/p75NTRsignaling in different aspects of adult neurogenesis. First, we observed that DG doublecortin (DCX) +progenitors express p75NTR both, in healthy humans and control animals, although the percentageof DCX+ cells are significantly reduced in AD. Interestingly, the expression of p75NTR in theseprogenitors is significantly decreased in AD conditions compared to controls. In order to assessthe contribution of the pro-NGF/p75NTR pathway to the memory deficits of APP/PS1 mice, weinjected pro-NGF neutralizing antibodies (anti-proNGF) into the DG of control and APP/PS1 miceand animals are subjected to a Morris water maze test. Intriguingly, we observed that anti-pro-NGF significantly restored memory performance of APP/PS1 animals and significantly increasethe percentage of DCX+ progenitors in the DG region of these animals. In summary, our resultssuggest that pro-NGF is involved in disrupting spatial memory in AD, at least in part by blockingadult neurogenesis. Moreover, we propose that adult neurogenesis alteration should be taken intoconsideration for better understanding of AD pathology. Additionally, we provide a new molecularentry point (pro-NGF/p75NTR signaling) as a promising therapeutic target in AD. This work was supported by “Fundació La Marató 2015” (C.E.). We thank, IRB Lleida Biobank (B.0000682), PLATAFORMA BIOBANCOS PT13/0010/0014, HUB-ICO-IDIBELL Biobankfor providing human tissue, and UAI IRBLleida for management support

Prostate cancer (PCa) in dogs is a highly malignant disease akin to its human counterpart. In contrast to the situation in humans, multi-gene approaches facilitating risk stratification of canine PCa are barely established. The aims of this study were the characterization of the transcriptional landscape of canine PCa and the identification of diagnostic, prognostic and/or therapeutic biomarkers through a multi-step screening approach. RNA-Sequencing of ten malignant tissues and fine-needle aspirations (FNA), and 14 nonmalignant tissues and FNAs was performed to find differentially expressed genes (DEGs) and deregulated pathways. The 4098 observed DEGs were involved in 49 pathways. These 49 pathways could be grouped into five superpathways summarizing the hallmarks of canine PCa: (i) inflammatory response and cytokines, (ii) regulation of the immune system and cell death, (iii) cell surface and PI3K signaling, (iv) cell cycle, and (v) phagosome and autophagy. Among the highly deregulated, moderately to strongly expressed DEGs that were members of one or more superpathways, 169 DEGs were listed in relevant databases and/or the literature and included members of the PCa pathway, oncogenes, prostate-specific genes, and druggable genes. These genes are novel and promising candidate diagnostic, prognostic and/or therapeutic canine PCa biomarkers.

Altres ajuts: Generalitat Valenciana APE/2021/022 One of the challenges of science in disease prevention is optimizing drug and vaccine delivery. Until now, many strategies have been employed in this sector, but most are quite complex and labile. To overcome these limitations, great efforts are directed to coupling drugs to carriers, either of natural or synthetic origin. Among the most studied cell carriers are antigen-presenting cells (APCs), however, red blood cells (RBCs) are positioned as attractive carriers in drug delivery due to their abundance and availability in the body. Furthermore, fish RBCs have a nucleus and have been shown to have a strong involvement in modulating the immune response. In this study, we evaluated the binding of three peptides to rainbow trout RBCs, two lectin-like peptides and another derived from Plasmodium falciparum membrane protein, in order to take advantage of this peptide-RBCs binding to generate tools to improve the specificity, efficacy, immunostimulatory ef-fect, and safety of the antiviral therapeutic or prophylactic administration systems currently used.

The protease activity in inflammatory bowel disease (IBD) and irritable bowel syndrome has been studied extensively using synthetic fluorogenic substrates targeting specific sets of proteases. We explored activities in colonic tissue from a 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis rat model by investigating the cleavage of bioactive peptides. Pure trypsin- and elastase-like proteases on the one hand and colonic tissue from rats with TNBS-induced colitis in the acute or post-inflammatory phase on the other, were incubated with relevant peptides to identify their cleavage pattern by mass spectrometry. An increased cleavage of several peptides was observed in the colon from acute colitis rats. The tethered ligand (TL) sequences of peptides mimicking the N-terminus of protease-activated receptors (PAR) 1 and 4 were significantly unmasked by acute colitis samples and these cleavages were positively correlated with thrombin activity. Increased cleavage of β-endorphin and disarming of the TL-sequence of the PAR3-based peptide were observed in acute colitis and linked to chymotrypsin-like activity. Increased processing of the enkephalins points to the involvement of proteases with specificities different from trypsin- or chymotrypsin-like enzymes. In conclusion, our results suggest thrombin, chymotrypsin-like proteases and a set of proteases with different specificities as potential therapeutic targets in IBD.

Neuroblastoma is a common extracranial solid tumour of childhood, responsible for 15% of cancer-related deaths in children. Prognoses vary from spontaneous remission to aggressive disease with extensive metastases, where treatment is challenging. Tumours are thought to arise from sympathoadrenal progenitor cells, which derive from an embryonic cell population called neural crest cells that give rise to diverse cell types, such as facial bone and cartilage, pigmented cells, and neurons. Tumours are found associated with mature derivatives of neural crest, such as the adrenal medulla or paraspinal ganglia. Sympathoadrenal progenitor cells express anaplastic lymphoma kinase (ALK), which encodes a tyrosine kinase receptor that is the most frequently mutated gene in neuroblastoma. Activating mutations in the kinase domain are common in both sporadic and familial cases. The oncogenic role of ALK has been extensively studied, but little is known about its physiological role. Recent studies have implicated ALK in neural crest migration and sympathetic neurogenesis. However, very few downstream targets of ALK have been identified. Here, we describe pathological activation of ALK in the neural crest, which promotes proliferation and migration, while preventing differentiation, thus inducing the onset of neuroblastoma. Understanding the effects of ALK activity on neural crest cells will help find new targets for neuroblastoma treatment.

Meniscus injury and meniscectomy are strongly related to osteoarthritis, thus there is a clinical need for meniscus replacement. The purpose of this study is to create a meniscus scaffold with micro-scale circumferential and radial fibres suitable for a one-stage cell-based treatment. Poly-caprolactone-based scaffolds with three different architectures were made using melt electrowriting (MEW) technology and their in vitro performance was compared with scaffolds made using fused-deposition modelling (FDM) and with the clinically used Collagen Meniscus Implants® (CMI®). The scaffolds were seeded with meniscus and mesenchymal stromal cells (MSCs) in fibrin gel and cultured for 28 d. A basal level of proteoglycan production was demonstrated in MEW scaffolds, the CMI®, and fibrin gel control, yet within the FDM scaffolds less proteoglycan production was observed. Compressive properties were assessed under uniaxial confined compression after 1 and 28 d of culture. The MEW scaffolds showed a higher Young’s modulus when compared to the CMI® scaffolds and a higher yield point compared to FDM scaffolds. This study demonstrates the feasibility of creating a wedge-shaped meniscus scaffold with MEW using medical-grade materials and seeding the scaffold with a clinically-feasible cell number and -type for potential translation as a one-stage treatment.

The rapid rise of multidrug-resistant (MDR) bacteria has once again caused bacterial infections to become a global health concern. Antimicrobial peptides (AMPs), also known as host defense peptides (HDPs), offer a viable solution to these pathogens due to their diverse mechanisms of actions, which include direct killing as well as immunomodulatory properties (e.g., anti-inflammatory activity). HDPs may hence provide a more robust treatment of bacterial infections. In this review, the advent of and the mechanisms that lead to antibiotic resistance will be described. HDP mechanisms of antibacterial and immunomodulatory action will be presented, with specific examples of how the HDP aurein 2.2 and a few of its derivatives, namely peptide 73 and cG4L73, function. Finally, resistance that may arise from a broader use of HDPs in a clinical setting and methods to improve biocompatibility will be briefly discussed.