Prioritization of cancer therapeutic targets using CRISPR-Cas9 screens.

Functional genomics approaches can overcome limitations-such as the lack of identification of robust targets and poor clinical efficacy-that hamper cancer drug development. Here we performed genome-scale CRISPR-Cas9 screens in 324 human cancer cell lines from 30 cancer types and developed a data-driven framework to prioritize candidates for cancer therapeutics. We integrated cell fitness effects with genomic biomarkers and target tractability for drug development to systematically prioritize new targets in defined tissues and genotypes. We verified one of our most promising dependencies, the Werner syndrome ATP-dependent helicase, as a synthetic lethal target in tumours from multiple cancer types with microsatellite instability. Our analysis provides a resource of cancer dependencies, generates a framework to prioritize cancer drug targets and suggests specific new targets. The principles described in this study can inform the initial stages of drug development by contributing to a new, diverse and more effective portfolio of cancer drug targets.

The pathogenicity of Plasmopara viticola: a review of evolutionary dynamics, infection strategies and effector molecules.

Oomycetes are filamentous organisms that resemble fungi in terms of morphology and life cycle, primarily due to convergent evolution. The success of pathogenic oomycetes lies in their ability to adapt and overcome host resistance, occasionally transitioning to new hosts. During plant infection, these organisms secrete effector proteins and other compounds during plant infection, as a molecular arsenal that contributes to their pathogenic success. Genomic sequencing, transcriptomic analysis, and proteomic studies have revealed highly diverse effector repertoires among different oomycete pathogens, highlighting their adaptability and evolution potential.The obligate biotrophic oomycete Plasmopara viticola affects grapevine plants (Vitis vinifera L.) causing the downy mildew disease, with significant economic impact. This disease is devastating in Europe, leading to substantial production losses. Even though Plasmopara viticola is a well-known pathogen, to date there are scarce reviews summarising pathogenicity, virulence, the genetics and molecular mechanisms of interaction with grapevine.This review aims to explore the current knowledge of the infection strategy, lifecycle, effector molecules, and pathogenicity of Plasmopara viticola. The recent sequencing of the Plasmopara viticola genome has provided new insights into understanding the infection strategies employed by this pathogen. Additionally, we will highlight the contributions of omics technologies in unravelling the ongoing evolution of this oomycete, including the first in-plant proteome analysis of the pathogen.

Deregulated immune cell recruitment orchestrated by c-MET impairs pulmonary inflammation and fibrosis.

BACKGROUND: Pulmonary fibrosis (PF) represents the pathologic end stage of several interstitial lung diseases (ILDs) associated with high morbidity and mortality rates. However, current treatments can only delay disease progression rather than provide a cure. The role of inflammation in PF progression is well-established, but new insights into immune regulation are fundamental for developing more efficient therapies. c-MET signaling has been implicated in the migratory capacity and effector functions of immune cells. Nevertheless, the role of this signaling pathway in the context of PF-associated lung diseases remains unexplored. METHODS: To determine the influence of c-MET in immune cells in the progression of pulmonary fibrosis, we used a conditional deletion of c-Met in immune cells. To induce pulmonary fibrosis mice were administered with bleomycin (BLM) intratracheally. Over the course of 21 days, mice were assessed for weight change, and after euthanasia at different timepoints, bronchoalveolar lavage fluid cells and lung tissue were assessed for inflammation and fibrosis. Furthermore, c-MET expression was assessed in cryobiopsy sections, bronchoalveolar lavage fluid cells samples and single cell RNA-sequencing dataset from human patients with distinct interstitial lung diseases. RESULTS: c-MET expression was induced in lung immune cells, specifically in T cells, interstitial macrophages, and neutrophils, during the inflammatory phase of BLM-induced PF mouse model. Deletion of c-Met in immune cells correlated with earlier weight recovery and improved survival of BLM-treated mice. Moreover, the deletion of c-Met in immune cells was associated with early recruitment of the immune cell populations, normally found to express c-MET, leading to a subsequent attenuation of the cytotoxic and proinflammatory environment. Consequently, the less extensive inflammatory response, possibly coupled with tissue repair, culminated in less exacerbated fibrotic lesions. Furthermore, c-MET expression was up-regulated in lung T cells from patients with fibrosing ILD, suggesting a potential involvement of c-MET in the development of fibrosing disease. CONCLUSIONS: These results highlight the critical contribution of c-MET signaling in immune cells to their enhanced uncontrolled recruitment and activation toward a proinflammatory and profibrotic phenotype, leading to the exacerbation of lung injury and consequent development of fibrosis.

Oligonucleotide probes for imaging and diagnosis of bacterial infections.

The rise of infectious diseases as a public health concern has necessitated the development of rapid and precise diagnostic methods. Imaging techniques like nuclear and optical imaging provide the ability to diagnose infectious diseases within the body, eliminating delays caused by sampling and pre-enrichments of clinical samples and offering spatial information that can aid in a more informed diagnosis. Traditional molecular probes are typically created to image infected tissue without accurately identifying the pathogen. In contrast, oligonucleotides can be tailored to target specific RNA sequences, allowing for the identification of pathogens, and even generating antibiotic susceptibility profiles by focusing on drug resistance genes. Despite the benefits that nucleic acid mimics (NAMs) have provided in terms of stabilizing oligonucleotides, the inadequate delivery of these relatively large molecules into the cytoplasm of bacteria remains a challenge for widespread use of this technology. This review summarizes the key advancements in the field of oligonucleotide probes for in vivo imaging, highlighting the most promising delivery systems described in the literature for developing optical imaging through in vivo hybridization.

The CD6 interactome orchestrates ligand-independent T cell inhibitory signaling.

BACKGROUND: T-cell membrane scaffold proteins are pivotal in T cell function, acting as versatile signaling hubs. While CD6 forms a large intracellular signalosome, it is distinguished from typical scaffolds like LAT or PAG by possessing a substantial ectodomain that binds CD166, a well-characterized ligand expressed on most antigen-presenting cells (APC), through the third domain (d3) of the extracellular region. Although the intact form of CD6 is the most abundant in T cells, an isoform lacking d3 (CD6∆d3) is transiently expressed on activated T cells. Still, the precise character of the signaling transduced by CD6, whether costimulatory or inhibitory, and the influence of its ectodomain on these activities are unclear. METHODS: We expressed CD6 variants with extracellular deletions or cytosolic mutations in Jurkat cells containing eGFP reporters for NF-κB and NF-AT transcription factor activation. Cell activation was assessed by eGFP flow cytometry following Jurkat cell engagement with superantigen-presenting Raji cells. Using imaging flow cytometry, we evaluated the impact of the CD6-CD166 pair on cell adhesiveness during the antigen-dependent and -independent priming of T cells. We also examined the role of extracellular or cytosolic sequences on CD6 translocation to the immunological synapse, using immunofluorescence-based imaging. RESULTS: Our investigation dissecting the functions of the extracellular and cytosolic regions of CD6 revealed that CD6 was trafficked to the immunological synapse and exerted tonic inhibition wholly dependent on its cytosolic tail. Surprisingly, however, translocation to the synapse occurred independently of the extracellular d3 and of engagement to CD166. On the other hand, CD6 binding to CD166 significantly increased T cell:APC adhesion. However, this activity was most evident in the absence of APC priming with superantigen, and thus, in the absence of TCR engagement. CONCLUSIONS: Our study identifies CD6 as a novel 'on/off' scaffold-receptor capable of modulating responsiveness in two ways. Firstly, and independently of ligand binding, it establishes signaling thresholds through tonic inhibition, functioning as a membrane-bound scaffold. Secondly, CD6 has the capacity for alternative splicing-dependent variable ligand engagement, modulating its checkpoint-like activity.

Socioeconomic inequalities in waiting times for breast cancer surgery.

Prompt access to cancer care is a policy priority in several OECD countries, because delayed access can exacerbate deleterious health outcomes. Access to care based on need remains a key pillar of publicly-funded health systems. This study tests for the presence of inequalities in waiting times by socioeconomic status for patients receiving breast cancer surgery (mastectomy or breast conserving surgery) in England using the Hospital Episode Statistics. We investigate separately the pre-COVID-19 period (April 2015-January 2020), and the COVID-19 period (February 2020-March 2022). We use linear regression models to study the association between waiting times and income deprivation measured at the patient's area of residence. We control for demographic factors, type and number of comorbidities, past emergency admissions and Healthcare Resource Groups, and supply-level factors through hospital fixed effects. In the pre-COVID-19 period, we do not find statistically significant associations between income deprivation in the patient's area of residence and waiting times for surgery. In the COVID-19 period, we find that patients living in the most deprived areas have longer waiting times by 0.7 days (given a mean waiting time of 20.6 days).

Mapping the Oxford Shoulder Score onto the EQ-5D utility index.

PURPOSE: In order to enable cost-utility analysis of shoulder pain conditions and treatments, this study aimed to develop and evaluate mapping algorithms to estimate the EQ-5D health index from the Oxford Shoulder Score (OSS) when health outcomes are only assessed with the OSS. METHODS: 5437 paired OSS and EQ-5D questionnaire responses from four national multicentre randomised controlled trials investigating different shoulder pathologies and treatments were split into training and testing samples. Separate EQ-5D-3L and EQ-5D-5L analyses were undertaken. Transfer to utility (TTU) regression (univariate linear, polynomial, spline, multivariable linear, two-part logistic-linear, tobit and adjusted limited dependent variable mixture models) and response mapping (ordered logistic regression and seemingly unrelated regression (SUR)) models were developed on the training sample. These were internally validated, and their performance evaluated on the testing sample. Model performance was evaluated over 100-fold repeated training-testing sample splits. RESULTS: For the EQ-5D-3L analysis, the multivariable linear and splines models had the lowest mean square error (MSE) of 0.0415. The SUR model had the lowest mean absolute error (MAE) of 0.136. Model performance was greatest in the mid-range and best health states, and lowest in poor health states. For the EQ-5D-5L analyses, the multivariable linear and splines models had the lowest MSE (0.0241-0.0278) while the SUR models had the lowest MAE (0.105-0.113). CONCLUSION: The developed models now allow accurate estimation of the EQ-5D health index when only the OSS responses are available as a measure of patient-reported health outcome.

Deep into the Apoplast: Grapevine and Plasmopara viticola Proteomes Reveal the Secret Beneath Host and Pathogen Communication at 6 h After Contact.

The apoplast is the first hub of plant-pathogen communication where pathogen effectors are recognized by plant defensive proteins and cell receptors, thus activating signal transduction pathways. As a result of this first contact, the host triggers a defense response that involves the modulation of extra- and intracellular proteins. In grapevine-pathogen interactions, little is known about the trafficking between extra- and intracellular spaces. Grapevine is an economically important crop that relies on heavy fungicide use to control several diseases, and a deeper knowledge on the activation of its immune response is crucial to define new control strategies. In this study, we focused on the first 6 h postinoculation with Plasmopara viticola to evaluate grapevine proteome modulation in the apoplast. The in planta P. viticola proteome was also assessed to enable a deeper understanding of plant-pathogen communication. Our results showed that several plant mechanisms are triggered in the tolerant grapevine cultivar Regent after inoculation, such as oomycete recognition, plant cell wall modifications, reactive oxygen species signaling, and secretion of proteins to disrupt oomycete structures. On the other hand, P. viticola proteins related to development and virulence were the most predominant. This pioneer study highlights the early dynamics of cellular communication in grapevine defense that leads to the successful establishment of an incompatible interaction.

Is myocardial fibrosis appropriately assessed by calibrated and 2D strain derived integrated backscatter?

AIMS: Increased collagen content of the myocardium modifies tissue reflectivity and integrated backscatter (IBS) indexes are suggested as markers of myocardial fibrosis (MF). We sought to assess the correlation between calibrated (c) IBS and bidimensional (2D) strain derived IBS with left ventricular (LV) MF in patients with severe aortic stenosis (AS). METHODS AND RESULTS: We made a prospective observational cohort study including 157 patients with severe AS referred for surgical aortic valve replacement (AVR), with complete preoperative transthoracic echocardiography, cardiac magnetic resonance (CMR) and endomyocardial biopsy (EMB) obtained from the anterior basal septum at the time of surgery. Two groups of 30 patients were specifically evaluated, with and without late gadolinium enhancement (LGE) at CMR. IBS was obtained at QRS peak from both parasternal long axis (PLAX) and apical-three-chamber (AP3C) views and measured in decibels (dB). Whole-cardiac cycle IBS at basal anterior septum was obtained from 2D longitudinal strain. Correlation analysis of reflectivity indexes was performed with global and segmental (anterior basal septum) values of native T1 and extracellular volume (ECV), and EMB collagen volume fraction (CVF) (Masson´s Trichrome). IBS values were compared in both group of patients (LGE + vs. LGE -). 60 patients (74 [36-74] years, 45% male) with high gradient (mean gradient: 63 ± 20mmHg), normal flow (45 ± 10mL/m2) AS and preserved left ventricular ejection fraction (60 ± 9%) were included. Basal septum cIBS was - 17.45 (-31.2-10.95) and - 9.17 ± 9.45dB from PLAX and A3C views, respectively. No significant correlations were found between IBS and both non-invasive CMR tissue characterization and CVF: median MF of 9.7(2.1-79.9)%. Acoustic indexes were not significantly different according to the presence of pre-operative LGE. CONCLUSION: In this group of patients with classical severe AS, IBS reflectivity indexes are of no added value to discriminate the presence of MF.

Fighting Methicillin-Resistant Staphylococcus aureus with Targeted Nanoparticles.

Antimicrobial resistance (AMR) is considered one of the greatest threats to global health. Methicillin-resistant Staphylococcus aureus (MRSA) remains at the core of this threat, accounting for about 90% of S. aureus infections widespread in the community and hospital settings. In recent years, the use of nanoparticles (NPs) has emerged as a promising strategy to treat MRSA infections. NPs can act directly as antibacterial agents via antibiotic-independent activity and/or serve as drug delivery systems (DDSs), releasing loaded antibiotics. Nonetheless, directing NPs to the infection site is fundamental for effective MRSA treatment so that highly concentrated therapeutic agents are delivered to the infection site while directly reducing the toxicity to healthy human cells. This leads to decreased AMR emergence and less disturbance of the individual's healthy microbiota. Hence, this review compiles and discusses the scientific evidence related to targeted NPs developed for MRSA treatment.

CD6-mediated inhibition of T cell activation via modulation of Ras.

BACKGROUND: CD6 is one of many cell surface receptors known to regulate signal transduction upon T cell activation. However, whether CD6 mediates costimulatory or inhibitory signals is controversial. When T cells engage with antigen presenting cells (APCs), CD6 interacts with its ligand CD166 at the cell-cell interface while the cytosolic tail assembles a complex signalosome composed of adaptors and effector enzymes, that may either trigger activating signaling cascades, or instead modulate the intensity of signaling. Except for a few cytosolic adaptors that connect different components of the CD6 signalosome, very little is known about the mechanistic effects of the cytosolic effectors that bind CD6. METHODS: Jurkat model T cells were transfected to express wild-type (WT) CD6, or a cytoplasmic truncation, signaling-disabled mutant, CD6Δcyt. The two resulting cell lines were directly activated by superantigen (sAg)-loaded Raji cells, used as APCs, to assess the net signaling function of CD6. The Jurkat cell lines were further adapted to express a FRET-based unimolecular HRas biosensor that reported the activity of this crucial GTPase at the immunological synapse. RESULTS: We show that deletion of the cytosolic tail of CD6 enhances T-cell responses, indicating that CD6 restrains T-cell activation. One component of the CD6-associated inhibitory apparatus was found to be the GTPase activating protein of Ras (RasGAP), that we show to associate with CD6 in a phosphorylation-dependent manner. The FRET HRas biosensor that we developed was demonstrated to be functional and reporting the activation of the T cell lines. This allowed to determine that the presence of the cytosolic tail of CD6 results in the down-regulation of HRas activity at the immunological synapse, implicating this fundamental GTPase as one of the targets inhibited by CD6. CONCLUSIONS: This study provides the first description of a mechanistic sequence of events underlying the CD6-mediated inhibition of T-cell activation, involving the modulation of the MAPK pathway at several steps, starting with the coupling of RasGAP to the CD6 signalosome, the repression of the activity of Ras, and culminating in the reduction of ERK1/2 phosphorylation and of the expression of the T-cell activation markers CD69 and IL-2R α chain. Video abstract.

Transcriptomic and methylation analysis of susceptible and tolerant grapevine genotypes following Plasmopara viticola infection.

Downy mildew, caused by the biotrophic oomycete Plasmopara viticola, is one of the most economically significant grapevine diseases worldwide. Current strategies to cope with this threat rely on the massive use of chemical compounds during each cultivation season. The economic costs and negative environmental impact associated with these applications increased the urge to search for sustainable strategies of disease control. Improved knowledge of plant mechanisms to counteract pathogen infection may allow the development of alternative strategies for plant protection. Epigenetic regulation, in particular DNA methylation, is emerging as a key factor in the context of plant-pathogen interactions associated with the expression modulation of defence genes. To improve our understanding of the genetic and epigenetic mechanisms underpinning grapevine response to P. viticola, we studied the modulation of both 5-mC methylation and gene expression at 6 and 24 h post-infection (hpi). Leaves of two table grape genotypes (Vitis vinifera), selected by breeding activities for their contrasting level of susceptibility to the pathogen, were analysed. Following pathogen infection, we found variations in the 5-mC methylation level and the gene expression profile. The results indicate a genotype-specific response to pathogen infection. The tolerant genotype (N23/018) at 6 hpi exhibits a lower methylation level compared to the susceptible one (N20/020), and it shows an early modulation (at 6 hpi) of defence and epigenetic-related genes during P. viticola infection. These data suggest that the timing of response is an important mechanism to efficiently counteract the pathogen attack.

The grapevine aspartic protease gene family: characterization and expression modulation in response to Plasmopara viticola.

MAIN CONCLUSION: Grapevine aspartic proteases gene family is characterized and five VviAPs appear to be involved in grapevine defense against downy mildew. Grapevine (Vitis vinifera L.) is one of the most important crops worldwide. However, it is highly susceptible to the downy mildew disease caused by Plasmopara viticola (Berk. & Curt.) Berl. & De Toni. To minimize the use of fungicides used to control P. viticola, it is essential to gain a deeper comprehension on this pathosystem and proteases have gained particular interest in the past decade. Proteases were shown to actively participate in plant-pathogen interactions, not only in the processes that lead to plant cell death, stress responses and protein processing/degradation but also as components of the recognition and signalling pathways. The aim of this study was to identify and characterize the aspartic proteases (APs) involvement in grapevine defense against P. viticola. A genome-wide search and bioinformatics characterization of the V. vinifera AP gene family was conducted and a total of 81 APs proteins, coded by 65 genes, were found. VviAPs proteins can be divided into three categories, similar to those previously described for other plants. Twelve APs coding genes were selected, and expression analysis was conducted at several time-points after inoculation in both compatible and incompatible interactions. Five grapevine APs may be involved in grapevine tolerance against P. viticola. Our findings provide an overall understanding of the VviAPs gene family and establish better groundwork to further describe the roles of VviAPs in defense against P. viticola.

Assessment of the Antibiofilm Performance of Chitosan-Based Surfaces in Marine Environments.

Marine biofouling is a natural process often associated with biofilm formation on submerged surfaces, creating a massive economic and ecological burden. Although several antifouling paints have been used to prevent biofouling, growing ecological concerns emphasize the need to develop new and environmentally friendly antifouling approaches such as bio-based coatings. Chitosan (CS) is a natural polymer that has been widely used due to its outstanding biological properties, including non-toxicity and antimicrobial activity. This work aims to produce and characterize poly (lactic acid) (PLA)-CS surfaces with CS of different molecular weight (Mw) at different concentrations for application in marine paints. Loligo opalescens pens, a waste from the fishery industry, were used as a CS source. The antimicrobial activity of the CS and CS-functionalized surfaces was assessed against Cobetia marina, a model proteobacterium for marine biofouling. Results demonstrate that CS targets the bacterial cell membrane, and PLA-CS surfaces were able to reduce the number of culturable cells up to 68% compared to control, with this activity dependent on CS Mw. The antifouling performance was corroborated by Optical Coherence Tomography since PLA-CS surfaces reduced the biofilm thickness by up to 36%, as well as the percentage and size of biofilm empty spaces. Overall, CS coatings showed to be a promising approach to reducing biofouling in marine environments mimicked in this work, contributing to the valorization of fishing waste and encouraging further research on this topic.

Two sides of the same story in grapevine-pathogen interactions.

Proteases are an integral part of plant defence systems, and their role in plant-pathogen interactions is unequivocal. Emerging evidence suggests that different protease families contribute to the establishment not only of hypersensitive response, priming, and signalling, but also of recognition events through complex proteolytic cascades. Moreover, they play a crucial role in pathogen/microbe-associated molecular pattern (PAMP/MAMP)-triggered immunity as well as in effector-triggered immunity. However, despite important advances in our understanding of the role of proteases in plant defence, the contribution of proteases to pathogen defence in grapevine remains poorly understood. In this review, we summarize current knowledge of the main grapevine pathosystems and explore the role of serine, cysteine, and aspartic proteases from both the host and pathogen point of views.

Composition and structure of the bryophyte community of Park Savanna in Marajó Island, Pará, Brazil.

Aiming to enrich the knowledge about the flora of savannas, this paper studied the composition and structure of the bryophyte community of Park Savanna areas in Marajó Island - PA. Biological material was collected within 60 100-m2 plots equally distributed in the dry season of 2016 and the rainy season of 2017 in five Park Savanna areas (SP-I to SP-V). The composition, density, richness and diversity of species and presence of indicator species were compared between the sampled areas and seasons. The species were classified according to the substrates colonized and ecological groups of light tolerance. Significant differences in SP-V indicated that the area was the main factor influencing the composition of bryophytes (p: 0.0001), with five indicator species. There were also significant differences in density (p = 0.0001168) and richness (p = 0.0001317) of bryophytes between seasons (p-value = 0.3393; p-value = 0.04065; p: 0.1081). There was a predominance of generalist (25 spp.) and corticolous (728 individuals) species, which were widely distributed in the sampled areas. Therefore, the structure of the bryophyte communities was not influenced by seasonality, and this indicates that these plants are adapted to the environmental conditions.

Development of Chitosan-Based Surfaces to Prevent Single- and Dual-Species Biofilms of Staphylococcus aureus and Pseudomonas aeruginosa.

Implantable medical devices (IMDs) are susceptible to microbial adhesion and biofilm formation, which lead to several clinical complications, including the occurrence of implant-associated infections. Polylactic acid (PLA) and its composites are currently used for the construction of IMDs. In addition, chitosan (CS) is a natural polymer that has been widely used in the medical field due to its antimicrobial and antibiofilm properties, which can be dependent on molecular weight (Mw). The present study aims to evaluate the performance of CS-based surfaces of different Mw to inhibit bacterial biofilm formation. For this purpose, CS-based surfaces were produced by dip-coating and the presence of CS and its derivatives onto PLA films, as well surface homogeneity were confirmed by contact angle measurements, Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). The antimicrobial activity of the functionalized surfaces was evaluated against single- and dual-species biofilms of Staphylococcus aureus and Pseudomonas aeruginosa. Chitosan-based surfaces were able to inhibit the development of single- and dual-species biofilms by reducing the number of total, viable, culturable, and viable but nonculturable cells up to 79%, 90%, 81%, and 96%, respectively, being their activity dependent on chitosan Mw. The effect of CS-based surfaces on the inhibition of biofilm formation was corroborated by biofilm structure analysis using confocal laser scanning microscopy (CLSM), which revealed a decrease in the biovolume and thickness of the biofilm formed on CS-based surfaces compared to PLA. Overall, these results support the potential of low Mw CS for coating polymeric devices such as IMDs where the two bacteria tested are common colonizers and reduce their biofilm formation.

Comparative in vitro study on the local tolerance and efficacy of benzalkonium chloride, myristalkonium chloride and nonoxynol-9 as active principles in vaginal contraceptives.

BACKGROUND: Spermicides have been identified as a potentially attractive alternative to hormonal contraceptives and/or intrauterine devices. Thus, this study aimed evaluating the efficacy and local tolerance of benzalkonium chloride (BKC) and myristalkonium chloride (MKC) contained in Pharmatex® vaginal formulations and compare them with nonoxynol-9 (N-9), the most common active ingredient in topical vaginal contraceptives. METHODS: Human normozoospermic samples were assessed for motility, viability, acrosome status and penetration ability after exposure to control, N-9 or different BKC and MKC doses for 0 and 10 minutes. Local tolerance on HeLa cells was evaluated by the Trypan-blue and MTT assays. RESULTS: Exposure to BKC and MKC reduced acrosome integrity while promoting total immobilisation and complete loss of sperm viability (p < .001, n = 15). Both compounds also compromised sperm penetration ability upon exposure (p < .001, n = 15). N-9 induced the same outcomes (p < .001, n = 15); nevertheless, it was more toxic to HeLa cells than BKC and MKC (p < .05, n = 14). CONCLUSIONS: BKC and MKC present strong in vitro spermicidal activity at lower doses than N-9 and were better tolerated after immediate exposure than N-9. Available Pharmatex® galenic formulations were as effective as products based on N-9.

Synthesis and biological effects of small molecule enhancers for improved recombinant protein production in plant cell cultures.

Histone deacetylases are involved in chromatin remodelling and thus play a vital role in the epigenetic regulation of gene expression. HDAC inhibitors alter the acetylation status of histone and non-histone proteins to regulate various cellular events such as transcription. Novel HDAC inhibitors were designed and synthesised to promote higher levels of recombinant protein production in tobacco cell cultures. The effect of these chemical enhancers on the epigenetic profiles in plant cells has been evaluated by molecular docking, in vitro and in vivo studies. The addition of these novel enhancers led to an increase in histone H3 acetylation levels that promoted an increase in the accumulation levels of the recombinant protein in cell culture. These results can pave the way for the application of these enhancers to improve the production of high value products in plant cell based systems.

Antibacterial Action Mechanisms of Honey: Physiological Effects of Avocado, Chestnut, and Polyfloral Honey upon Staphylococcus aureus and Escherichia coli.

Numerous studies have explored the antibacterial properties of different types of honey from all around the world. However, the data available describing how honey acts against bacteria are few. The aim of this study was to apply a flow cytometry (FC) protocol to examine and characterize the primary effects of three varieties of honey (avocado, chestnut and polyfloral) upon physiological status of Staphylococcus aureus and Escherichia coli cells to reveal their antibacterial action mechanisms. The effects of honey samples on membrane potential, membrane integrity, and metabolic activity were assessed using different fluorochromes, in a 180 min time course assay. Time-kill experiments were also carried out under similar conditions. Exposure of S. aureus and E. coli to the distinct honey samples resulted in physiological changes related to membrane polarization and membrane integrity. Moreover, honey induced a remarkable metabolic disruption as primary physiological effect upon S. aureus. The different honey samples induced quite similar effects on both bacteria. However, the depth of bacteria response throughout the treatment varied depending on the concentration tested and among honey varieties, probably due to compositional differences in the honey.