Rhizobacterial diversity of Portuguese olive cultivars in the Douro valley and their potential as plant growth promoters.

AIMS: This study investigated the bacterial communities in the rhizosphere of two traditional Portuguese olive cultivars, Cobrançosa and Negrinha de Freixo, in relation to soil properties. Additionally, we aimed to isolate and identify bacteria with potential for biocontrol and other plant growth-promoting traits from these rhizosphere communities. METHODS AND RESULTS: Bacterial communities in the olive rhizosphere were investigated using a metabarcoding approach and the soil physicochemical properties of the olive groves were also analyzed. Higher bacterial richness was associated with Negrinha de Freixo growing in soil with high organic matter content and water-holding capacity. In contrast, the soils of the Cobrançosa grove presented higher pH and electric conductivity. Negrinha de Freixo rhizosphere was enriched with ASVs (Amplicon Sequence Variants) belonging to Bacillus, Gaiella, Acidothermus, Bradyrhizobium, and uncultured Xanthobacteraceae. On the other hand, the Cobrançosa rhizosphere was characterized by higher relative abundance of Streptomyces and Sphingomonas. Bacterial isolation from the rhizosphere and screening for plant growth-promoting activities were also performed. Six bacteria strains, predominantly Bacillus isolated from Negrinha de Freixo, demonstrated antagonistic activities against the olive fungal pathogen Colletotrichum gloeosporoides and other plant growth promotion (PGP) traits. CONCLUSIONS: Our findings demonstrate that the structure of rhizosphere bacterial communities associated with olive trees is shaped by both plant cultivar and soil-related factors. The higher number of bacterial species in the rhizosphere of Negrinha de Freixo was related to a higher organic matter content and a greater abundance of isolates with plant growth promotion traits, particularly Bacillus strains.

Multiplex Determination of Glycan Profiles on Urinary Prostate-Specific Antigen by Quartz-Crystal Microbalance Combined with Surface-Enhanced Raman Scattering.

Prostate cancer remains a major health concern, with prostate-specific antigen (PSA) being a key biomarker for its detection and monitoring. However, PSA levels often fall into a "gray zone", where PSA levels are not clearly indicative of cancer, thus complicating early diagnosis and treatment decisions. Glycosylation profiles, which often differ between healthy and diseased cells, have emerged as potential biomarkers to enhance the specificity and sensitivity of cancer diagnosis in these ambiguous cases. We propose the integration of two complementary techniques, namely quartz-crystal microbalance with dissipation (QCM-D) and surface-enhanced Raman scattering (SERS) to study PSA glycan profiles. QCM-D offers real-time operation, PSA mass quantification, and label-free detection with high sensitivity, as well as enhanced specificity and reduced cross-reactivity when using nucleic acid aptamers as capture ligands. Complementary SERS sensing enables the determination of the glycosylation pattern on PSA, at low concentrations and without the drawbacks of photobleaching, thereby facilitating multiplexed glycosylation pattern analysis. This integrated setup could retrieve a data set comprising analyte concentrations and associated glycan profiles in relevant biological samples, which may eventually improve early disease detection and monitoring. Prostate-specific antigen (PSA), a glycoprotein secreted by prostate epithelial cells, serves as our proof-of-concept analyte. Our platform allows multiplex targeting of PSA multiplex glycosylation profiles of PSA at "gray zone" concentrations for prostate cancer diagnosis. We additionally show the use of SERS for glycan analysis in PSA secreted from prostate cancer cell lines after androgen-based treatment. Differences in PSA glycan profiles from resistant cell lines after androgen-based treatment may eventually improve cancer treatment.

Engineering the Rhizosphere Microbiome with Plant Growth Promoting Bacteria for Modulation of the Plant Metabolome.

Plant-growth-promoting bacteria (PGPB) have beneficial effects on plants. They can promote growth and enhance plant defense against abiotic stress and disease, and these effects are associated with changes in the plant metabolite profile. The research problem addressed in this study was the impact of inoculation with PGPB on the metabolite profile of Salicornia europaea L. across controlled and field conditions. Salicornia europaea seeds, inoculated with Brevibacterium casei EB3 and Pseudomonas oryzihabitans RL18, were grown in controlled laboratory experiments and in a natural field setting. The metabolite composition of the aboveground tissues was analyzed using GC-MS and UHPLC-MS. PGPB inoculation promoted a reconfiguration in plant metabolism in both environments. Under controlled laboratory conditions, inoculation contributed to increased biomass production and the reinforcement of immune responses by significantly increasing the levels of unsaturated fatty acids, sugars, citric acid, acetic acid, chlorogenic acids, and quercetin. In field conditions, the inoculated plants exhibited a distinct phytochemical profile, with increased glucose, fructose, and phenolic compounds, especially hydroxybenzoic acid, quercetin, and apigenin, alongside decreased unsaturated fatty acids, suggesting higher stress levels. The metabolic response shifted from growth enhancement to stress resistance in the latter context. As a common pattern to both laboratory and field conditions, biopriming induced metabolic reprogramming towards the expression of apigenin, quercetin, formononetin, caffeic acid, and caffeoylquinic acid, metabolites that enhance the plant's tolerance to abiotic and biotic stress. This study unveils the intricate metabolic adaptations of Salicornia europaea under controlled and field conditions, highlighting PGPB's potential to redesign the metabolite profile of the plant. Elevated-stress-related metabolites may fortify plant defense mechanisms, laying the groundwork for stress-resistant crop development through PGPB-based inoculants, especially in saline agriculture.

Examining nutrition strategies to influence DNA methylation and epigenetic clocks: a systematic review of clinical trials.

Nutrition has powerful impacts on our health and longevity. One of the mechanisms by which nutrition might influence our health is by inducing epigenetic modifications, modulating the molecular mechanisms that regulate aging. Observational studies have provided evidence of a relationship between nutrition and differences in DNA methylation. However, these studies are limited in that they might not provide an accurate control of the interactions between different nutrients, or between nutrition and other lifestyle behaviors. Here we systematically reviewed clinical studies examining the impact of nutrition strategies on DNA methylation. We examined clinical studies in community-dwelling adults testing the effects of nutrition interventions on i) global DNA methylation and its proxies, and ii) epigenetic clocks. We included 21 intervention studies that focused on the effects of healthy nutrition patterns, specific foods or nutrients, as well as the effect of multivitamin or multimineral supplements. In four studies on the methylation effects of healthy dietary patterns, as defined by being rich in vegetables, fruits, whole-grains, and nuts and reduced in the intake of added sugars, saturated fat, and alcohol, two of them suggested that a healthy diet, is associated with lower epigenetic age acceleration, one of them reported increases in global DNA methylation, while another one found no diet effects. Studies examining epigenetic effects of specific foods, nutrients, or mixtures of nutrients were scarce. For both folic acid and polyunsaturated fatty acids, the available independent studies produced conflicting findings. Although more evidence is still needed to draw firm conclusions, results begin to suggest that healthy dietary patterns have positive effects on DNA methylation. Additional evidence from large randomized-controlled clinical trials is needed to support the effects of healthy nutrition on the DNA methylome.

Epithelial tubule interconnection driven by HGF-Met signaling in the kidney.

The formation of functional epithelial tubules is a central feature of many organ systems. Although the process of tubule formation by epithelial cells is well-studied, the way in which tubules connect with each other (i.e. anastomose) to form functional networks both in vivo and in vitro is not well understood. A key, unanswered question in the kidney is how the renal vesicles of the embryonic kidney connect with the nascent collecting ducts to form a continuous urinary system. We performed a ligand-receptor pair analysis on single cell RNA-seq data from embryonic mouse kidney tubules undergoing anastomosis to select candidates that might mediate this process in vivo. This analysis identified hepatocyte growth factor (HGF), which has known roles in cell proliferation, migration, and tubulogenesis, as one of several possible candidates. To test this possibility, we designed a novel assay to quantitatively examine epithelial tubule anastomosis in vitro using epithelial spheroids with fluorescently-tagged apical surfaces to enable direct visualization of anastomosis. This revealed that HGF is a potent inducer of tubule anastomosis. Tubule anastomosis occurs through a proliferation-independent mechanism that acts through the MAPK signaling cascade and matrix metalloproteinases (MMPs), the latter suggestive of a role in extracellular matrix turnover. Accordingly, treatment of explanted embryonic mouse kidneys with HGF and collagenase was sufficient to induce kidney tubule anastomosis. These results lay the groundwork for investigating how to promote functional interconnections between tubular epithelia, which have important clinical implications for utilizing in vitro grown kidney tissue in transplant medicine.

Plasma levels of adipokines and insulin are associated with markers of brain atrophy and cognitive decline in the spectrum of Alzheimer's Disease.

The discovery that metabolic alterations often coexist with neurodegenerative conditions has sparked interest in the examination of metabolic regulatory factors as potential modulators of brain health. Here, we examined the role of adipokines (leptin, adiponectin, resistin, and IL6) and insulin on different markers of brain atrophy in participants on the spectrum of Alzheimer's Disease. We included 566 participants from the Alzheimer's Disease Neuroimaging Initiative (ADNI) dataset with 1063 follow-up time points (average follow-up: one year); and examined the association between metabolic regulatory factors and volumetric MRI values, white matter hyperintensities, and measures of cognitive impairment. Higher leptin, resistin, IL6, and insulin were associated with markers of cerebral atrophy, such as lower total brain volume, or higher ventricular volume. Higher leptin and resistin were also associated with greater impairment in daily life activities. Higher adiponectin was associated with lower ventricle volume. There was no association between adipokines or insulin with white matter hyperintensities. Our findings indicate a co-occurrence between alterations in metabolic regulatory factors and in brain volume along the preclinical to clinical spectrum of Alzheimer's Disease. These results suggest that strategies aimed at promoting metabolic health may positively impact brain health.

Timing of P2Y12 Inhibitor Administration in Patients With STEMI Undergoing Primary PCI.

BACKGROUND: The optimal timing of P2Y12 inhibitor administration in patients with ST-segment elevation myocardial infarction (STEMI) has not been completely elucidating. OBJECTIVES: This analysis from a prospective multicenter registry sought to assess the safety and effectiveness of P2Y12 inhibitor pretreatment in patients transferred for primary percutaneous coronary intervention (PCI) within a regional STEMI network. METHODS: Pretreatment was defined as P2Y12 inhibitor administration before coronary angiography. Endpoints were major adverse cardiac events (MACE), major bleeding, and net adverse clinical events, a composite of MACE or major bleeding, within 30 days of index admission. Association of P2Y12 inhibitor pretreatment with outcomes was modeled using doubly robust weighted estimators based on propensity score analysis. RESULTS: Of 1,624 patients included, 1,033 received P2Y12 inhibitors before angiography and 591 in the catheterization laboratory (cath lab). The non-pretreated cohort more often had history of coronary artery disease and were more likely to receive antiplatelet therapy before the index admission. After adjustment for confounding and dependent censoring, pretreatment with P2Y12 inhibitors predicted lower risk of MACE (adjusted HR: 0.53; 95% CI: 0.37-0.76), without increasing bleeding risk (adjusted HR: 0.62; 95% CI: 0.36-1.05), resulting in superior net clinical benefit (adjusted HR: 0.47; 95% CI: 0.26-0.86) compared with in-cath lab administration of P2Y12 inhibitors. There was a significant treatment-by-time interaction for MACE risk, whereby the observed benefits of pretreatment only became apparent when time between P2Y12 inhibitor administration and PCI was longer than 80 minutes. CONCLUSIONS: In contemporary patients with STEMI transferred for primary PCI, pretreatment with P2Y12 inhibitors was associated with a significant time-dependent reduction of 30-day MACE without increasing bleeding risk.

Outcomes of transvaginal radiofrequency ablation for symptomatic leiomyomas.

OBJECTIVE: Describe the effect of transvaginal radiofrequency ablation for leiomyoma in symptomatic patients and post procedure follow-up. MATERIAL AND METHODS: A retrospective forward-looking observational study was performed including 63 patients who underwent transvaginal radiofrequency ablation between January 2016 and December 2022 at San Cecilio University Hospital in Granada, Spain. The variables registered were: age, parity, the clinical features that lead to the medical visit and pre-surgical treatment. Prior to the procedure, leiomyoma location and volume were determined by transvaginal ultrasound. Follow-ups were scheduled at 6 and 12 months to evaluate symptom improvement, adverse outcomes, leiomyoma volume and if any necessary post-surgical treatment was required. RESULTS: Mean leiomyoma volume at baseline, 6 months and 12 months was 83.3 (24.9-130.7), 42.4 (4.7-89.0) and 19.2 (1.9-80.4) cm3, respectively (p < .001). Significant differences were found between the baseline and 12 month visits (p < .001). At the annual follow-up, the mean rate of volume reduction was 79.5 %, being higher in women who reported symptom improvement compared to those who reported no change in symptom intensity from baseline (84.6% vs. 30.8 %). Patients with a lower initial volume and age over 40 were more likely to have treatment efficacy. 8 pregnancies were registered post procedure. CONCLUSION: Radiofrequency is well tolerated, allowing for same-day discharge, rapid recovery and a safe approach for women who want to preserve their reproductive potential. Initial volume and age over 40 appear to be factors that should be considered in patient selection. Further studies are needed to continue evaluating the outcomes and identifying predictive factors.

Prevalence of Gastrointestinal Parasites in Small Ruminant Farms in Southern Spain.

The primary population of small ruminants in Spain is concentrated in the southern region, a critical area for the country's livestock production. Indirect economic losses can occur when this livestock is affected by gastrointestinal parasites. This study aimed to determine the prevalence of these parasites in small ruminant herds (159 sheep and 39 goats) through coprological analyses and conducted a survey on farmers' management practices related to gastrointestinal parasite control. The survey results revealed some important aspects: monitoring through coprological analyses is not a common practice; veterinarians are not typically involved in deworming plans; anthelmintic treatment in adults is often applied twice a year in sheep and once a year in goats; and finally, drug rotation was higher in sheep farms. Coprological analyses showed Eimeria spp. as the most common parasitic infection, followed by Strongyles infection. Other parasites like Moniezia spp., Trichuris spp., and D. dendriticum were less important, although their prevalence was higher in sheep than goats. This constitutes the first report on the epidemiological status of gastrointestinal parasites in small ruminants in southern Spain. Based on the survey findings, the introduction of certain management measures on farms could potentially mitigate parasite infections.

Prevalence and Risk Factors for Colonization by Multidrug-Resistant Microorganisms among Long-Term Travelers and Recently Arrived Migrants.

Multidrug-resistant (MDR) bacteria have become one of the most important health problems. We aimed to assess whether international travel may facilitate their spread through the colonization of asymptomatic travelers. A cross-sectional study was conducted (November 2018 to February 2022). Pharyngeal and rectal swabs were obtained from long-term travelers and recently arrived migrants from non-European countries, and an epidemiological survey was performed. Colonization by Gram-negative bacteria and methicillin-resistant Staphylococcus aureus (MRSA) was determined by chromogenic media and MALDI-TOF-MS. Resistance mechanisms were determined by the biochip-based molecular biology technique. Risk factors for colonization were assessed by logistic regression. In total, 122 participants were included: 59 (48.4%) recently arrived migrants and 63 (51.6%) long-term travelers. After their trip, 14 (11.5%) participants-5 (8.5%) migrants and 9 (14.3%) travelers-had rectal colonization by one MDR bacterium. Escherichia coli carrying the extended-spectrum beta-lactamase (ESBL) CTX-M-15 was the most frequent. No participants were colonized by MRSA or carbapenemase-producing Enterobacteriaceae. The only risk factor independently associated with MDR bacterial colonization was previous hospital attention [OR, 95% CI: 10.16 (2.06-50.06)]. The risk of colonization by MDR bacteria among recently arrived migrants and long-term travelers is similar in both groups and independently associated with previous hospital attention.

[Translated article] Syringes for intraocular administration: A systematic review.

OBJECTIVE: The off-label use in clinical practice of non-approved syringes for intravitreal drug administration has resulted in the detection of silicone oil drops in the vitreous of some patients. This situation derives from the lack of approved syringes for intraocular use in the Spanish market. The aim of this work is to review the use of syringes for intraocular administration, as well as to search for alternatives that meet the legal requirements for these unmet needs. METHOD: A systematic review was performed following the PRISMA 2020 guidelines by searching PubMed with the descriptors: (silicone) AND (syringes) AND ((intraocular) OR (intravitreal)) and filtering all existing publications from January 2006 to December 2023, including all those articles dealing with silicone oil release in intravitreal injections and analysing the possible consequences. RESULTS: Sixty-eight results were found, 23 of which were excluded because they did not deal with the subject under study, leaving a total of 45 articles for the systematic review. These were classified according to the conclusions obtained in 4 groups: the adverse reactions produced by silicone; the administration technique; the physicochemical aspects of silicone release; and the characteristics of the medical device. After reviewing the current manufacturers and technical data sheets of commercialised syringes, the existing syringes for this use have been collected, finding 2 that will probably be commercialised in Spain at the beginning of 2024: Zero Residual™ 0.2 ml SiO-free and VitreJect® Ophthalmic. CONCLUSIONS: From the results obtained, it can be interpreted that the use of syringes and needles with silicone for intravitreal use is a concern for health professionals due to the implications and consequences that may arise in patients, the most important being adverse reactions, so it is necessary to have silicone-free syringes on the market that are specific for intraocular use. Safety and legality in the use of intraocular syringes and needles is essential to guarantee ocular integrity and patient health.

Safety Profile and Effectiveness of Dapagliflozin in Pediatric Patients with Chronic Kidney Disease.

INTRODUCTION: Nephroprotection in pediatric chronic kidney disease (CKD) has a major positive impact, both on residual renal function and on quality of life, by delaying the need for renal replacement therapy. To this day, nephroprotective drugs used in children are mainly limited to angiotensin-converting enzyme (ACE) inhibitors and angiotensin-receptor blockers; interestingly, as suggested by trials conducted in adults with CKD, sodium-glucose cotransporter 2 inhibitors (SGLT2i) might also be beneficial to pediatric patients. However, there are no validated data to this date documenting the effect of SGLT2i in pediatric patients with CKD. METHODS: We present a retrospective single-center study reporting the use of dapagliflozin in pediatric patients with CKD, aiming to evaluate dapagliflozin safety profile as well as its potential for renal protection. Our study describes 7 patients with a mean age of 13.3 years (+/- 7.029) presenting with identified glomerulopathy, leading to CKD and already treated by ACE inhibitors. Patients received a daily dose of dapagliflozin of 5 or 10 mg. RESULTS: Over a period of 15 months, all patients reported the medication as easy to use. After an initial dip, estimated glomerular filtration rate decline slope stabilized in all patients. Urinary albumin-over-creatinine ratio had a strong tendency to decrease after 6 months of treatment (p = 0.0684). Systolic blood pressure also had a tendency to decrease after 6 months of treatment (p = 0.1). No significant side effect was reported by the patients. CONCLUSION: The promising results presented in this study support the use of SGLT2i in pediatric patients with CKD, although larger, randomized controlled trials in pediatric patients are necessary to better characterize their effectiveness in this particular population.

[Pharmacological intervention in the medication review of institutionalised elderly patients under polypharmacy]. / Intervención farmacéutica en la revisión del tratamiento en pacientes mayores polimedicados institucionalizados.

OBJECTIVE: To describe the Drug-Related Problems (DRP) and their resolution after pharmacological review in institutionalised elderly patients under polypharmacy. DESIGN: Descriptive, retrospective cohort study from January to October of 2022. LOCATION: Twelve nursing homes at the Community of Madrid. PARTICIPANTS: 295 patients aged 65 or older taking at least 5 chronic medications prescribed prior to the treatment review. INTERVENTIONS: Medication reviews carried out by the pharmacist and agreed upon in face-to-face meetings between the primary care doctor, the nursing home doctor and the pharmacist. MAIN MEASUREMENTS: Detected DRP, types and resolution. A age, sex, and number of medications before and after the intervention. Pharmacological subgroups according to anatomical therapeutic chemical classification system (ATC) and active pharmaceutical ingredients involved in the detected DRPs. RESULTS: 1425 DRP were detected, with a mean of 4.85 (SD 3.33) DRPs/patient. The most frequent DRP was reconciliation error (32.52%), followed by pharmaceutical regimen and dosaje. Among the 1425 improvement proposals, 86.73% of them were accepted.Significant statistically differences were observed between the number of medications per patient prior to the pharmacotherapy review (12.29) and after it (10.20), obtaining an average difference of 2.09 (95%CI: 1.98-2.21; P<.001). CONCLUSIONS: It is found that the intervention of multidisciplinary team in which the pharmacist performs a revision of the medication decreased the number of prescribed medications. Therefore, it reduces polymedication and its associated risks.

Mortality risk factors in the dependent population of Castilla-La Mancha (Spain) before and during the first COVID-19 wave.

The coronavirus disease pandemic has had an important impact worldwide. The population aged over 65 years and aged dependent persons are the population groups which have suffered in a highest level the consequences of the pandemic in terms of cases and death. In Spain, the situation is similar to other countries, but regional studies are needed because competencies on long-term care depend on regional public administration. Thus, the aim of this work is to analyse social and individual factors associated with the risk of mortality of legally recognised dependent people during the pandemic compared to a non-pandemic period. The data were extracted from the administrative database on individuals included in Castilla-La Mancha's long-term care system and it was merged with the information from the Spanish National Death Index administered by the Ministry of Health, Consumption and Social Welfare. The results show that the risk of mortality between March and June 2020 was positively associated with being male; being older than 65, with an especially high impact in the group aged over 90; having a higher level of dependency; living in a nursing home; and living in a place with more population density. Intraregional differences related to health areas also exists in both pandemic and non-pandemic periods. These findings are critical with a view to enhancing protocols for the care of the most vulnerable population groups.

Transcription-coupled repair of DNA-protein cross-links depends on CSA and CSB.

Covalent DNA-protein cross-links (DPCs) are toxic DNA lesions that block replication and require repair by multiple pathways. Whether transcription blockage contributes to the toxicity of DPCs and how cells respond when RNA polymerases stall at DPCs is unknown. Here we find that DPC formation arrests transcription and induces ubiquitylation and degradation of RNA polymerase II. Using genetic screens and a method for the genome-wide mapping of DNA-protein adducts, DPC sequencing, we discover that Cockayne syndrome (CS) proteins CSB and CSA provide resistance to DPC-inducing agents by promoting DPC repair in actively transcribed genes. Consequently, CSB- or CSA-deficient cells fail to efficiently restart transcription after induction of DPCs. In contrast, nucleotide excision repair factors that act downstream of CSB and CSA at ultraviolet light-induced DNA lesions are dispensable. Our study describes a transcription-coupled DPC repair pathway and suggests that defects in this pathway may contribute to the unique neurological features of CS.

Development of a Green, Quick, and Efficient Method Based on Ultrasound-Assisted Extraction Followed by HPLC-DAD for the Analysis of Bioactive Glycoalkaloids in Potato Peel Waste.

α-Solanine and α-chaconine are the two most predominant glycoalkaloids (GAs) present in potato. Potato peel contains a high concentration of GAs, which are especially interesting for application in the pharmaceutical industry due to their different beneficial properties (such as anticarcinogenic, anti-inflammatory, antiallergic, antipyretic, antiviral, fungicide, and antibiotic activities, among others); so, potato peel waste can be valorized by extracting these biologically active compounds. For this, a green, quick, and efficient miniaturized analytical approach based on ultrasound-assisted extraction (UAE) combined with HPLC-DAD was developed to quantify α-solanine and α-chaconine in potato peel. Some parameters of the extraction were optimized, including the extraction method, the type of solvent, and the sample/solvent ratio, by a three-factor, three-level (33) full factorial experimental design. The optimal extraction conditions were obtained with UAE using methanol as a solvent and a sample/solvent ratio of 1:10 (w/v, g/mL). The analytical greenness metric for sample preparation (AGREEprep) tool was used to assess the greenness of the methods used. The tool revealed an acceptable green analysis, with 0.61 points. The method was validated and applied to the evaluation of GAs in the peel of 15 commercial varieties of potato. The amount of glycoalkaloids found in the samples evaluated ranged from 143 to 1273 mg/kg and from 117 to 1742 mg/kg dry weight for α-solanine and α-chaconine, respectively. These results reveal the important variability that exists between potato varieties; so, their analysis is of great importance to select the most suitable ones for biovalorization (e.g., the Amandine and Rudolph varieties, with around 3000 mg/kg, in total, of both GAs). To provide higher stability to the peel during storage, freeze-drying or a medium-temperature drying process resulted preferable to avoid GA degradation. Overall, this study will contribute to the expansion of the future biovalorization of potato peel waste as well as provide a powerful analytical tool for GA analysis.

CRISPR Activation in Mouse Trophoblast Stem Cells.

The placenta is a vital organ that regulates nutrient supply to the developing embryo during gestation. In mice, the placenta is composed of trophoblast lineage and mesodermal derivatives, which merge through the chorioallantoic fusion process in a critical event for the progression of placenta development. The trophoblast lineage is derived from self-renewing, multipotent cells known as mouse trophoblast stem cells (mTSCs). These cells are a valuable tool that allows scientists to comprehend the signals regulating major placental cell types' self-renewal and differentiation capacity. Recent advances in CRISPR-Cas9 genome editing applied in mTSCs have provided novel insights into the molecular networks involved in placentation. Here, we present a comprehensive CRISPR activation (CRISPRa) protocol based on the CRISPR/gRNA-directed synergistic activation mediator (SAM) method to overexpress specific target genes in mTSCs.

Syringes for intraocular administration: Systematic review. / Jeringas para administración intraocular: revisión sistemática.

OBJECTIVE: The off-label use in clinical practice of non-approved syringes for intravitreal drug administration has resulted in the detection of silicone oil drops in the vitreous of some patients. This situation derives from the lack of approved syringes for intraocular use in the Spanish market. The aim of this work is to review the use of syringes for intraocular administration, as well as to search for alternatives that meet the legal requirements for these unmet needs. METHOD: A systematic review was performed following the PRISMA 2020 Guidelines by searching PubMed with the descriptors: "silicone" AND "syringes" AND ("intraocular" OR "intravitreal") and filtering all existing publications from January 2006 to December 2023, including all those articles dealing with silicone oil release in intravitreal injections and analysing the possible consequences. RESULTS: Sixty-eight results were found, 23 of which were excluded because they did not deal with the subject under study, leaving a total of 45 articles for the systematic review. These were classified according to the conclusions obtained in 4 groups: the adverse reactions produced by silicone, the administration technique, the physicochemical aspects of silicone release, and the characteristics of the medical device. After reviewing the current manufacturers and technical data sheets of commercialized syringes, the existing syringes for this use have been collected, finding two that will probably be commercialized in Spain at the beginning of 2024: Zero Residual™ 0.2 ml SiO-free and VitreJect® Ophthalmic. CONCLUSIONS: From the results obtained, it can be interpreted that the use of syringes and needles with silicone for intravitreal use is a concern for health professionals due to the implications and consequences that may arise in patients, the most important being adverse reactions, so it is necessary to have silicone-free syringes on the market that are specific for intraocular use. Safety and legality in the use of intraocular syringes and needles is essential to guarantee ocular integrity and patient health.

G-quadruplex DNA structure is a positive regulator of MYC transcription.

DNA structure can regulate genome function. Four-stranded DNA G-quadruplex (G4) structures have been implicated in transcriptional regulation; however, previous studies have not directly addressed the role of an individual G4 within its endogenous cellular context. Using CRISPR to genetically abrogate endogenous G4 structure folding, we directly interrogate the G4 found within the upstream regulatory region of the critical human MYC oncogene. G4 loss leads to suppression of MYC transcription from the P1 promoter that is mediated by the deposition of a de novo nucleosome alongside alterations in RNA polymerase recruitment. We also show that replacement of the endogenous MYC G4 with a different G4 structure from the KRAS oncogene restores G4 folding and MYC transcription. Moreover, we demonstrate that the MYC G4 structure itself, rather than its sequence, recruits transcription factors and histone modifiers. Overall, our work establishes that G4 structures are important features of transcriptional regulation that coordinate recruitment of key chromatin proteins and the transcriptional machinery through interactions with DNA secondary structure, rather than primary sequence.

Reduction of Chronic Primary Low Back Pain by Spinal Manipulative Therapy is Accompanied by Decreases in Segmental Mechanical Hyperalgesia and Pain Catastrophizing: A Randomized Placebo-controlled Dual-blind Mixed Experimental Trial.

Chronic primary low back pain (CPLBP) refers to low back pain that persists over 3 months, that cannot be explained by another chronic condition, and that is associated with emotional distress and disability. Previous studies have shown that spinal manipulative therapy (SMT) is effective in relieving CPLBP, but the underlying mechanisms remain elusive. This randomized placebo-controlled dual-blind mixed experimental trial (NCT05162924) aimed to investigate the efficacy of SMT to improve CPLBP and its underlying mechanisms. Ninety-eight individuals with CPLBP and 49 controls were recruited. Individuals with CPLBP received SMT (n = 49) or a control intervention (n = 49), 12 times over 4 weeks. The primary outcomes were CPLBP intensity (0-100 on a numerical rating scale) and disability (Oswestry Disability Index). Secondary outcomes included pressure pain thresholds in 4 body regions, pain catastrophizing, Central Sensitization Inventory, depressive symptoms, and anxiety scores. Individuals with CPLBP showed widespread mechanical hyperalgesia (P < .001) and higher scores for all questionnaires (P < .001). SMT reduced pain intensity compared with the control intervention (mean difference: -11.7 [95% confidence interval, -11.0 to -12.5], P = .01), but not disability (P = .5). Similar mild to moderate adverse events were reported in both groups. Mechanical hyperalgesia at the manipulated segment was reduced after SMT compared with the control intervention (P < .05). Pain catastrophizing was reduced after SMT compared with the control intervention (P < .05), but this effect was not significant after accounting for changes in clinical pain. Although the reduction of segmental mechanical hyperalgesia likely contributes to the clinical benefits of SMT, the role of pain catastrophizing remains to be clarified. PERSPECTIVE: This randomized controlled trial found that 12 sessions of SMT yield greater relief of CPLBP than a control intervention. These clinical effects were independent of expectations, and accompanied by an attenuation of hyperalgesia in the targeted segment and a modulation of pain catastrophizing.