In recent years, Rwanda, especially its Eastern Province, has been contending with water shortages, primarily due to prolonged dry spells and restricted water sources. This situation poses a substantial threat to the country’s agriculture-based economy and food security. The impact may escalate with climate change, exacerbating the frequency and severity of droughts. However, there is a lack of comprehensive spatiotemporal analysis of meteorological and agricultural droughts, which is an urgent need for a nationwide assessment of the drought’s impact on vegetation and agriculture. Therefore, the study aimed to identify meteorological and agricultural droughts by employing the Standardized Precipitation Evapotranspiration Index (SPEI) and the Vegetation Health Index (VHI). VHI comprises the Vegetation Condition Index (VCI) and the Temperature Condition Index (TCI), both derived from the Normalized Difference Vegetation Index (NDVI) and Land Surface Temperature (LST). This study analyzed data from 31 meteorological stations spanning from 1983 to 2020, as well as remote sensing indices from 2001 to 2020, to assess the spatiotemporal patterns, characteristics, and adverse impact of droughts on vegetation and agriculture. The results showed that the years 2003, 2004, 2005, 2006, 2013, 2014, 2015, 2016, and 2017 were the most prolonged and severe for both meteorological and agricultural droughts, especially in the Southern Province and Eastern Province. These extremely dry conditions led to a decline in both vegetation and crop production in the country. It is recommended that policymakers engage in proactive drought mitigation activities, address climate change, and enforce water resource management policies in Rwanda. These actions are crucial to decreasing the risk of drought and its negative impact on both vegetation and crop production in Rwanda. Full article

The wavefront reconstruction is a crucial step in determining the performance of wavefront detection instruments. The wavefront reconstruction algorithm is primarily evaluated in three dimensions: accuracy, speed, and noise immunity. In this paper, we propose a hybrid zonal reconstruction algorithm that introduces slope and curvature information in the diagonal, anti-diagonal, horizontal, and vertical directions by dividing the neighbor sampling points into subregions in groups of four. By canceling the same parameters in integration equations, an algorithm using multi-directional slope–curvature information is achieved with only two sets of integration equations in each subregion, reducing the processing time. Simulation experiments show that the relative root-mean-square reconstruction error of this algorithm is improved by about 4 orders of magnitude compared with existing algorithms that use multi-directional slope information or slope–curvature information alone. Compared with the hybrid multi-directional slope–curvature algorithm, the proposed algorithm can reduce computation time by about 50% as well as provide better noise immunity and reconstruction accuracy. Finally, the validity of the proposed algorithm is verified by the null test experiment. Full article

In tunnel boring projects, wear and tear in the tooling system can have significant consequences, such as decreased boring efficiency, heightened maintenance costs, and potential safety hazards. In this paper, a fault diagnosis method for TBM tooling systems based on SAV−SVDD failure location (SSFL) is proposed. The aim of this method is to detect faults caused by disk cutter wear during the boring process, which diminishes the boring efficiency and is challenging to detect during construction. This paper uses SolidWorks to create a complete three−dimensional model of the TBM hydraulic thrust system and tool system. Then, dynamic simulations are performed with Adams. This helps us understand how the load on the propulsion hydraulic cylinder changes as the TBM tunneling tool wears to different degrees during construction. The hydraulic propulsion system was modeled and simulated using AMESIM software. Utilizing the load on the hydraulic propulsion cylinder as an input signal, pressure signals from the two chambers of the hydraulic cylinder and the system’s flow signal were acquired. This enabled an in−depth exploration of the correlation between these acquired signals and the extent of the tooling system failure. Following this analysis, a collection of normal sample data and sample data representing different degrees of disk cutter abrasions was amassed for further study. Next, an SSFL network model for locating the failure area of the cutter was established. Fault sample data were used as the input, and the accuracy of the fault diagnosis model was tested. The test results show that the performance of the SSFL network model is better than that of the SAE−SVM and SVDD network models. The SSFL model achieves 90% accuracy in determining the failure area of the cutter head. The model effectively identifies the failure regions, enabling timely tool replacement to avoid decreased boring efficiency under wear conditions. The experimental findings validate the feasibility of this approach. Full article

Interleukin (IL)-32 is produced by T lymphocytes, natural killer cells, monocytes, and epithelial cells. IL-32 induces the production of pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α, IL-1β, IL-6, and IL-8, and IL-32 expression is highly increased in rheumatoid arthritis (RA) patients. Enolase-1 (ENO1) is a glycolytic enzyme and the stimulation of ENO1 induces high levels of pro-inflammatory cytokines in concanavalin A (Con A)-activated peripheral blood mononuclear cells (PBMCs) and macrophages in RA patients. In addition, there are many reports that anti-ENO1 antibody is correlated with the disease progression of RA. It implies that ENO1 could regulate IL-32 production during inflammation related to the pathogenesis of RA. Therefore, we investigated the role of ENO1 in IL-32 production using Con A-activated PBMCs and RA PBMCs. IL-32 expression is increased by ENO1 stimulation using real-time PCR and ELISA. In addition, we confirmed that IL-32 production was decreased in Con A-activated PBMCs and RA PBMCs pre-treated with NF-κB or p38 MAPK pathway inhibitors. Taken together, these results suggest that ENO1 plays an important role in inflammation through the induction of IL-32 production by the activation of the NF-κB and p38 MAPK pathways. Full article

Worldwide, oak species are threatened with extinction due to habitat loss, pathogens, and changing fire regimes. Ex situ conservation through tissue culture may protect the remaining genetic diversity of Quercus dumosa, or the coastal sage scrub oak, from further loss. We designed three basal salt formulations based on the mineral composition of shoot tips and first leaves from mature Q. dumosa and explored carbohydrate source, stress-mitigating compounds, and plant growth regulator concentrations to develop a method of cultivating many Q. dumosa culture lines in vitro. All three novel basal salt formulations led to decreased necrosis compared with commercial basal salt formulas WPM, MS, and DKW. Substitution of 30 g L⁻¹ sucrose with glucose and adding 250 mg L⁻¹ ascorbic acid, 5.2 mg L⁻¹ SNP sodium nitroprusside, and 103 mg L⁻¹ y-aminobutyric acid improved culture health overall. In an experiment involving 115 culture lines, 0.66 mg L⁻¹ 6-benzylaminopurine produced the highest average shoots per explant, but 0.33 mg L⁻¹ produced the greatest proportion of shoots 2 cm or greater. Incubation for 24 h in 20 mg L⁻¹ indole-3-butyric acid led to the most rooting. These methods show promise for the ex situ conservation of many genotypes of endangered Q. dumosa. Full article

Cascade genetic testing and surveillance reduce morbidity and mortality in Lynch syndrome. However, barriers to conveying information about genetic disorders within families result in low uptake of genetic testing. Provider-mediated interventions may increase uptake but raise legal and ethical concerns. We describe 30 years of national experience with cascade genetic testing combining family- and provider-mediated contact in Lynch syndrome families in the Danish Hereditary Non-Polyposis Colorectal Cancer (HNPCC) Register. We aimed to estimate the added value of information letters to family members in Lynch syndrome families (provider-mediated contact) compared to family members not receiving such letters and thus relying on family-mediated contact. National clinical practice for cascade genetic testing, encompassing infrastructure, legislation, acceptance, and management of the information letters, is also discussed. Cascade genetic testing resulted in 7.3 additional tests per family. Uptake of genetic testing was 54.4% after family-mediated and 64.9% after provider-mediated contact, corresponding to an odds ratio of 1.8 (p < 0.001). The uptake of genetic testing was highest in the first year after diagnosis of Lynch syndrome in the family, with 72.5% tested after provider-mediated contact. In conclusion, the Danish model combining family- and provider-mediated contact can increase the effect of cascade genetic testing. Full article

Over 3000 iron formation caves (IFCs) have formed in erosion-resistant Fe(III)-rich rocks throughout Brazil. Microbial Fe(III) reduction occurs in IFCs, where a microbe-rich, Fe(III)-depleted paste (sub muros) is found behind an Fe(III)-(hydr)oxide crust in the ceiling/walls. Microbial Fe(III) reduction in sub muros appears to be responsible for the transformation of Fe(III) to more soluble Fe(II), which is removed, leading to cave formation. This process of biospeleogenesis is likely controlled by O₂ availability, which is linked to seasonal changes. Here, we studied the effects of alternating anoxia/oxia on the microbial community and on Fe solubility in banded iron formation (BIF), a rock type consisting of layered Fe(III)-oxide and silicate. Incubations of synthetic pore water, pulverized BIF, and sub muros were prepared and incubated under anoxia, during which BIF-Fe(III) reduction proceeded. During the Fe(III) reduction period, Firmicutes and/or Alphaproteobacteria were enriched, and genes involved in Fe(III) and sulfate reduction were detected in the metagenomes. Fe(II) oxidation genes, which were detected in the fresh sub muros, were not found. Upon the addition of atmospheric O₂, Fe(III) reduction was arrested, and incomplete Fe(II) oxidation occurred. Betaproteobacteria, Gammaproteobacteria, and Chloroflexi increased in relative abundance following aeration, and Fe(III) reduction genes were still identified. Our results demonstrate that the sub muros microbial community retains the ability to reduce Fe(III) and drive speleogenesis despite fluctuations in O₂ levels. Full article

Menopausal status affects the prognoses and consequences of breast cancer. Therefore, this retrospective study aimed to reveal the molecular variation profile differences in breast cancer patients according to their menopausal status, with the hypothesis that the molecular variation profiles will be different at premenopausal and postmenopausal ages. Breast cancer patients (n = 254) who underwent molecular subtyping and QIAseq Human Breast Cancer NGS Panel screening between 2018 and 2022 were evaluated retrospectively. Their menopausal status was defined by age, and those aged 50 years and above were considered postmenopausal. Of the subjects, 58.66% (n = 149) were premenopausal and 41.34% (n = 105) were postmenopausal. The mean age at the time of diagnosis for all patients was 49.31 ± 11.19 years, with respective values of 42.11 ± 5.51 and 59.54 ± 9.01 years for the premenopausal and postmenopausal groups, respectively (p = 0.000). Among premenopausal patients, the percentages of patients in BCa subtypes (luminal A, luminal B-HER2(−), luminal B-HER2(+), HER2 positive, and triple-negative) were determined to be 34.90%, 8.05%, 26.17%, 10.74%, and 20.13%, respectively, while in the postmenopausal group, these values were 39.05%, 16.19%, 24.76%, 6.67%, and 13.33%, respectively (p > 0.05). Considering menopausal status, the distribution of hormone receptors in premenopausal patients was ER(+)/PgR(+) 63.76%, ER(−)/PgR(−) 23.49%, ER(+)/PgR(−) 10.74%, and ER(−)/PgR(+) 2.01%, respectively, while in postmenopausal women, this distribution was observed to be 74.29%, 23.81%, 1.90% and 0.00% in the same order (p = 0.008). The most frequently mutated gene was TP53 in 130 patients (51.18%), followed by PIK3CA in 85 patients (33.46%), BRCA2 and NF1 in 56 patients (22.05%), PTEN in 54 patients (21.26%), and ATR and CHEK2 in 53 patients (20.87%). TP53, PIK3CA, NF1, BRCA2, PTEN, and CHEK2 mutations were more frequently observed in premenopausal patients, while TP53, PIK3CA, BRCA2, BRCA1, and ATR mutations in postmenopausal patients. These findings contribute to a deeper understanding of the underlying causes of breast cancer with respect to menopausal status. This study is the first from Turkey that reflects the molecular subtyping and somatic mutation profiles of breast cancer patients according to menopausal status. Full article

Frequency-domain near-infrared spectroscopy (FD-NIRS) has been used for non-invasive assessment of cortical oxygenation since the late 1990s. However, there is limited research demonstrating clinical validity and general reproducibility. To address this limitation, recording duration for adequate validity and within- and between-day reproducibility of prefrontal cortical oxygenation was evaluated. To assess validity, a reverse analysis of 10-min-long measurements (n = 52) at different recording durations (1–10-min) was quantified via coefficients of variation and Bland–Altman plots. To assess within- and between-day within-subject reproducibility, participants (n = 15) completed 2-min measurements twice a day (morning/afternoon) for five consecutive days. While 1-min recordings demonstrated sufficient validity for the assessment of oxygen saturation (S_tO₂) and total hemoglobin concentration (THb), recordings ≥4 min revealed greater clinical utility for oxy- (HbO) and deoxyhemoglobin (HHb) concentration. Females had lower S_tO_2, THb, HbO, and HHb values than males, but variability was approximately equal between sexes. Intraclass correlation coefficients ranged from 0.50–0.96. The minimal detectable change for S_tO₂ was 1.15% (95% CI: 0.336–1.96%) and 3.12 µM for THb (95% CI: 0.915–5.33 µM) for females and 2.75% (95%CI: 0.807–4.70%) for S_tO₂ and 5.51 µM (95%CI: 1.62–9.42 µM) for THb in males. Overall, FD-NIRS demonstrated good levels of between-day reliability. These findings support the application of FD-NIRS in field-based settings and indicate a recording duration of 1 min allows for valid measures; however, data recordings of ≥4 min are recommended when feasible. Full article

The increase in the size and complexity of large knowledge graphs now available online has resulted in the emergence of many approaches focusing on enabling the quick exploration of the content of those data sources. Structural non-quotient semantic summaries have been proposed in this direction that involve first selecting the most important nodes and then linking them, trying to extract the most useful subgraph out of the original graph. However, the current state of the art systems use costly centrality measures for identifying the most important nodes, whereas even costlier procedures have been devised for linking the selected nodes. In this paper, we address both those deficiencies by first exploiting embeddings for node selection, and then by meticulously selecting approximate algorithms for node linking. Experiments performed over two real-world big KGs demonstrate that the summaries constructed using our method enjoy better quality. Specifically, the coverage scores obtained were 0.8, 0.81, and 0.81 for DBpedia v3.9 and 0.94 for Wikidata dump 2018, across 20%, 25%, and 30% summary sizes, respectively. Additionally, our method can compute orders of magnitude faster than the state of the art. Full article

Citric acid cycle deficiencies are extremely rare due to their central role in energy metabolism. The ACO2 gene encodes the mitochondrial isoform of aconitase (aconitase 2), the second enzyme of the citric acid cycle. Approximately 100 patients with aconitase 2 deficiency have been reported with a variety of symptoms, including intellectual disability, hypotonia, optic nerve atrophy, cortical atrophy, cerebellar atrophy, and seizures. In this study, a homozygous deletion in the ACO2 gene in two brothers with reduced aconitase 2 activity in fibroblasts has been described with symptoms including truncal hypotonia, optic atrophy, hyperopia, astigmatism, and cerebellar atrophy. In an in vivo trial, triheptanoin was used to bypass the defective aconitase 2 and fill up the citric acid cycle. Motor abilities in both patients improved. Full article

Mediterranean marine caves have been categorized as both biodiversity reservoirs and vulnerable habitats. However, only a few studies have focused on Porifera assemblages within marine caves along the Adriatic Apulian coast (southern Italy). In this study, the sponge fauna of the Rondinella cave, a semi-submerged marine cave along the coast of Bari (Southern Adriatic Sea), was investigated for the first time. The use of advanced image analysis in combination with targeted sampling has made it possible to determine the spatial distribution and diversity of Porifera along a transect from the entrance to the end of the cave. Data analysis clustered the stations into two groups, separated according to the distance from the entrance and corresponding to the cave entrance and the semi-dark zone. Sponges were found at all stations covering a considerable part of the substrate, with the highest cover values occurring in the semi-dark zone. A total of 54 sponge taxa were identified: 49 Demospongiae, 3 Homoscleromorpha, and 2 Calcarea. Six species are new records for the Apulian marine caves, one species represents a new record for marine caves, and two species are new findings for the southern Adriatic Sea. Full article

Implementing best management practices (BMPs) has proven to be an efficient method for reducing non-point source (NPS) pollutants. Agricultural NPS pollution is considered to be a major contributor to water quality impairment. This study aims to assess the variation in hydrologic and water quality outputs at field and watershed scales when BMPs are implemented using modeling approaches. The Yazoo River Watershed (YRW) is the largest watershed basin in the state of Mississippi with approximately 50% agricultural land. Runoff generated from agricultural areas carries sediments and nutrients. The Merigold watershed (MW) is a sub-basin of the YRW and a field-scale watershed with most of the land use being agriculture. It is essential to quantify the streamflow, sediment, total nitrogen (TN), and total phosphorus (TP) when BMPs are implemented. BMPs such as vegetative filter strips (VFS) and cover crops (CC) were tested in this study. The Soil and Water Assessment Tool (SWAT) model was applied to quantify the watershed’s hydrologic and water quality outputs. SWAT model accuracy assessment was performed by calibration and validation process using the Nash and Sutcliffe Efficiency Index (NSE). Model performance was satisfactory for monthly streamflow, with NSE values in the range of 0.62 to 0.81, and for daily sediments, TN, and TP load estimation, with NSE values of 0.21, 0.20, and 0.47, respectively. CC was planted after harvesting the main crop. Therefore, it is essential to quantify the seasonal reduction in pollutants. Water quality was improved after BMP implementation, and an overall decrease in streamflow, sediment, TN, and TP loads was observed for both MW and YRW during dry and wet seasons. Previous studies regarding seasonal assessments with CC implementation in the MW and YRW were limited. Therefore, the results from this study could be a unique addition to the scientific literature. Full article

Major depressive disorder is a severe mood disorder associated with a marked decrease in quality of life and social functioning, accompanied by a risk of suicidal behavior. Therefore, seeking out and adhering to effective treatment is of great personal and society-wide importance. Weight changes associated with antidepressant therapy are often cited as the reason for treatment withdrawal and thus are an important topic of interest. There indeed exists a significant mechanistic overlap between depression, antidepressant treatment, and the regulation of appetite and body weight. The suggested pathomechanisms include the abnormal functioning of the homeostatic (mostly humoral) and hedonic (mostly dopaminergic) circuits of appetite regulation, as well as causing neuromorphological and neurophysiological changes underlying the development of depressive disorder. However, this issue is still extensively discussed. This review aims to summarize mechanisms linked to depression and antidepressant therapy in the context of weight change. Full article

In the context of AI, as algorithms rapidly penetrate e-commerce platforms, it is timely to investigate the role of algorithm awareness (AA) in privacy decisions because it can shape consumers’ information-disclosure behaviors. Focusing on the role of AA in the privacy decision-making process, this study investigated consumers’ personal information disclosures when using an e-commerce platform with personalized algorithms. By integrating the dual calculus model and the theory of planned behavior (TPB), we constructed a privacy decision-making model for consumers. Sample data from 581 online-shopping consumers were collected by a questionnaire survey, and SmartPLS 4.0 software was used to conduct a structural equation path analysis and a mediating effects test on the sample data. The findings suggest that AA is a potential antecedent to the privacy decision-making process through which consumers seek to evaluate privacy risks and make self-disclosure decisions. The privacy decision process goes through two interrelated trade-offs—that threat appraisals and coping appraisals weigh each other to determine the (net) perceived risk and, then, the (net) perceived risk and the perceived benefit weigh each other to decide privacy attitudes. By applying the TPB to the model, the findings further show that privacy attitudes and subjective norms jointly affect information-disclosure intention whereas perceived behavioral control has no significant impact on information-disclosure intention. The results of this study give actionable insights into how to utilize the privacy decision-making process to promote algorithm adoption and decisions regarding information disclosure, serving as a point of reference for the development of a human-centered algorithm based on AA in reference to FEAT. Full article

During periodontitis, the extracellular capsule of Porphyromonas gingivalis favors alveolar bone loss by inducing Th1 and Th17 patterns of lymphocyte response in the infected periodontium. Dendritic cells recognize bacterial antigens and present them to T lymphocytes, defining their activation and polarization. Thus, dendritic cells could be involved in the Th1 and Th17 response induced against the P. gingivalis capsule. Herein, monocyte-derived dendritic cells were obtained from healthy individuals and then stimulated with different encapsulated strains of P. gingivalis or two non-encapsulated isogenic mutants. Dendritic cell differentiation and maturation were analyzed by flow cytometry. The mRNA expression levels for distinct Th1-, Th17-, or T-regulatory-related cytokines and transcription factors, as well as TLR2 and TLR4, were assessed by qPCR. In addition, the production of IL-1β, IL-6, IL-23, and TNF-α was analyzed by ELISA. The encapsulated strains and non-encapsulated mutants of P. gingivalis induced dendritic cell maturation to a similar extent; however, the pattern of dendritic cell response was different. In particular, the encapsulated strains of P. gingivalis induced higher expression of IRF4 and NOTCH2 and production of IL-1β, IL-6, IL-23, and TNF-α compared with the non-encapsulated mutants, and thus, they showed an increased capacity to trigger Th1 and Th17-type responses in human dendritic cells. Full article

The vanadium redox flow battery (VRFB) is a highly favorable tool for storing renewable energy, and the catalytic activity of electrode materials is crucial for its development. Taurine-functionalized carbon nanotubes (CNTs) were prepared with the aim of augmenting the redox process of vanadium ions and enhancing the efficiency of the VRFB. Sulfonated CNTs were synthesized through a simple modification process in a taurine solution and used as electrocatalysts for redox reactions involving VO²⁺/VO₂⁺ and V²⁺/V³⁺. The SO₃H-CNTs modified at 60 °C for 2 h exhibit the best electrocatalytic activity, showing higher redox peak current values compared to pristine carboxylated CNTs (COOH-CNTs). Sulfonic acid groups added to the surface of CNTs increase active sites for redox reactions and act as carriers for mass transfer and bridges for charge transfer, accelerating the rate of the electrode reactions. A battery consisting of SO₃H-CNTs as catalysts demonstrates the outstanding charge–discharge performance at a current density of 300 mA∙cm⁻². This configuration displays voltage and energy efficiencies of 81.46% and 78.83%, respectively, representing enhancements of 6.15% and 6.12% compared to that equipped with conventional graphite felts (75.31%, 72.71%). This study illustrates that taurine-functionalized carbon nanotubes serve as an efficient and promising catalyst for both the anode and cathode, leading to the improved performance of the VRFB. Full article

In this paper, we provide a novel methodology for high-precision positioning that utilizes 1-bit additional information, which applies to various positioning techniques. The proposed approach leverages binary information to indicate if a user is within a specified space of interest and refines the estimated location information outside this area. By matching the estimated locations outside the area of interest with the valid location information within, this methodology corrects the positional data obtained through any arbitrary positioning technique, aligning the estimated positions with the intended spatial boundaries. Performance analysis metrics, such as Average Positioning Error (APE) and Cumulative Distribution Function for positioning coverage, were employed to assess the effectiveness of the proposed methods. Numerical simulations demonstrate how the proposed method enhances the averaged positioning accuracy, significantly outperforming the conventional time of arrival method. Furthermore, the proposed positioning correction methodology demonstrates validated feasibility applicable to an arbitrary existing positioning method. Full article

Given the global challenge of increasing teacher attrition and turnover rates, the exploration of factors and mechanisms that improve teachers’ organizational commitment has become a pivotal topic in educational research. In this context, the present study examines the influence of teachers’ emotional intelligence on their organizational commitment, with a specific inquiry into the mediating role of teachers’ psychological well-being and the moderating role of principal transformational leadership, as informed by the broaden-and-build theory of positive emotions and the trait activation theory. We verified this study’s hypotheses based on 768 valid questionnaires collected from Chinese primary and secondary school teachers. The results reveal that teachers’ emotional intelligence can predict their organizational commitment both directly and indirectly through the mediating role of psychological well-being. Additionally, principal transformational leadership amplifies the positive effect of teachers’ emotional intelligence on psychological well-being and, subsequently, organizational commitment. These findings theoretically deepen our understanding of the psychological pathways and the boundary conditions linking teachers’ emotional intelligence to their organizational commitment, while also offering valuable practical implications for building a stable and effective teaching workforce. Full article

When pipe defects are generated above the T-type support structure location, it is difficult to distinguish the reflection signals caused by the weld bead at the support structure from the reflection echoes of pipe defects. Therefore, in order to effectively detect pipe defects, a waveform subtraction method with a circumferential shear horizontal (CSH) guided wave is proposed, which is generated by an electromagnetic acoustic transducer (EMAT). First, a CSH₀ guided wave mode with a center frequency of 500 kHz is selected to establish a three-dimensional model with and without pipe defects above the support structure. Following this, the influence of different widths of support structures on the echo signal is compared. Moreover, simulation and experimental results are used to compare the influence of different welding qualities on the detection results. Finally, the waveform subtraction method is used to process the simulation and experimental signals, and the influence of pipe defects with different lengths and depths is discussed. The results show that the non-through crack defect of 5 mm × 1 mm (length × depth) can be detected. The results show that this method can effectively detect the cracks by eliminating the influence of the weld echo, which provides a new concept for the detection of the defect above the support structure. Full article

Lipid nanoparticles (LNPs) have established their position as nonviral vectors for gene therapy. Tremendous efforts have been made to modulate the properties of LNPs to unleash their full clinical potential. Among the strategies being pursued, the layer-by-layer (LbL) technique has gained considerable attention in the biomedical field. Illuminated by our previous work, here we investigate if the LbL approach could be used to modify the LNP cores formulated with three different ionizable lipids: DODMA, MC3, and DODAP. Additionally, we wondered if more than three layers could be loaded onto LNPs without disrupting their gene transfection ability. Taking advantage of physicochemical analysis, as well as uptake and gene silencing studies, we demonstrate the feasibility of modifying the surface of LNPs with the LbL assembly. Precisely, we successfully modified three different LNPs using the layer-by-layer strategy which abrogated luciferase activity in vitro. Additionally, we constructed a 5×-layered HA-LNP containing the MC3 ionizable lipid which outperformed the 3×-layered counterpart in transfecting miRNA-181-5p to the pediatric GBM cell line, as a proof-of-concept in vitro experiment. The method used herein has been proven reproducible, of easy modification to adapt to different ionizable lipid-containing LNPs, and holds great potential for the translation of RNA-based therapeutic strategies. Full article

Porphyromonas gingivalis (Pg) and its gingipain proteases contribute to Alzheimer’s disease (AD) pathogenesis through yet unclear mechanisms. Cellular secretion of small extracellular vesicles or exosomes (EXO) increases with aging as part of the senescence-associated secretory phenotype (SASP). We have shown that EXO isolated from Pg-infected dendritic cells contain gingipains and other Pg antigens and transmit senescence to bystander gingival cells, inducing alveolar bone loss in mice in vivo. Here, EXO were isolated from the gingiva of mice and humans with/without periodontitis (PD) to determine their ability to penetrate the blood–brain barrier (BBB) in vitro and in vivo. PD was induced by Pg oral gavage for 6 weeks in C57B6 mice. EXO isolated from the gingiva or brain of donor Pg-infected (PD EXO) or control animals (Con EXO) were characterized by NTA, Western blot, and TEM. Gingival PD EXO or Con EXO were labeled and injected into the gingiva of uninfected WT mouse model. EXO biodistribution in brains was tracked by an in vivo imaging system (IVIS) and confocal microscopy. The effect of human PD EXO on BBB integrity and permeability was examined using TEER and FITC dextran assays in a human in vitro 3D model of the BBB. Pg antigens (RGP and Mfa-1) were detected in EXO derived from gingival and brain tissues of donor Pg-infected mice. Orally injected PD EXO from donor mice penetrated the brains of recipient uninfected mice and colocalized with hippocampal microglial cells. IL-1β and IL-6 were expressed in human PD EXO and not in Con EXO. Human PD EXO promoted BBB permeability and penetrated the BBB in vitro. This is the first demonstration that microbial-induced EXO in the oral cavity can disseminate, cross the BBB, and may contribute to AD pathogenesis. Full article