Improvement of Phased Antenna Array Applied in Focused Microwave Breast Hyperthermia.

Focused microwave breast hyperthermia (FMBH) employs a phased antenna array to perform beamforming that can focus microwave energy at targeted breast tumors. Selective heating of the tumor endows the hyperthermia treatment with high accuracy and low side effects. The effect of FMBH is highly dependent on the applied phased antenna array. This work investigates the effect of polarizations of antenna elements on the microwave-focusing results by simulations. We explore two kinds of antenna arrays with the same number of elements using different digital realistic human breast phantoms. The first array has all the elements' polarization in the vertical plane of the breast, while the second array has half of the elements' polarization in the vertical plane and the other half in the transverse plane, i.e., cross polarization. In total, 96 sets of different simulations are performed, and the results show that the second array leads to a better focusing effect in dense breasts than the first array. This work is very meaningful for the potential improvement of the antenna array for FMBH, which is of great significance for the future clinical applications of FMBH. The antenna array with cross polarization can also be applied in microwave imaging and sensing for biomedical applications.

Validation of mitochondrial biomarkers and immune dynamics in polycystic ovary syndrome.

PROBLEM: Polycystic ovary syndrome (PCOS), a prevalent endocrine-metabolic disorder, presents considerable therapeutic challenges due to its complex and elusive pathophysiology. METHOD OF STUDY: We employed three machine learning algorithms to identify potential biomarkers within a training dataset, comprising GSE138518, GSE155489, and GSE193123. The diagnostic accuracy of these biomarkers was rigorously evaluated using a validation dataset using area under the curve (AUC) metrics. Further validation in clinical samples was conducted using PCR and immunofluorescence techniques. Additionally, we investigate the complex interplay among immune cells in PCOS using CIBERSORT to uncover the relationships between the identified biomarkers and various immune cell types. RESULTS: Our analysis identified ACSS2, LPIN1, and NR4A1 as key mitochondria-related biomarkers associated with PCOS. A notable difference was observed in the immune microenvironment between PCOS patients and healthy controls. In particular, LPIN1 exhibited a positive correlation with resting mast cells, whereas NR4A1 demonstrated a negative correlation with monocytes in PCOS patients. CONCLUSION: ACSS2, LPIN1, and NR4A1 emerge as PCOS-related diagnostic biomarkers and potential intervention targets, opening new avenues for the diagnosis and management of PCOS.

Changes in physicochemical and gut microbiota fermentation property induced by acetylation of polysaccharides from Cyperus esculentus.

In this study, the impact of acetylation on physicochemical, digestive behavior and fermentation characteristics of Cyperus esculentus polysaccharides (CEP) was investigated. Results indicated that the acetylation led the molecules to be more likely aggregated, followed by a higher crystallinity, a lower apparent viscosity and a higher ratio of G" to G' (tan δ). Importantly, the acetylated polysaccharides (ACEP) had a lower digestibility, but its molecular weight was lower than that of original polysaccharides (CEP) following a simulated saliva-gastrointestinal digestion. Gut microbiota fermentation indicated that both polysaccharides generated outstanding short-chain fatty acids (SCFAs), in which the acetylated polysaccharides had a faster fermentation kinetics than the original one, followed by a quicker reduction of pH and a more accumulation of SCFAs, particularly butyrate. Fermentation of both polysaccharides promoted Akkermansia, followed by a reduced richness of Klebsiella. Importantly, the current study revealed that the fermentation of acetylated polysaccharides enriched Parabacteroides, while fermentation of original ones promoted Bifidobacterium, for indicating their individual fermentation characteristics and gut environmental benefits.

Uncovering the potential of APOD as a biomarker in gastric cancer: A retrospective and multi-center study.

Gastric cancer (GC) poses a significant health challenge worldwide, necessitating the identification of predictive biomarkers to improve prognosis. Dysregulated lipid metabolism is a well-recognized hallmark of tumorigenesis, prompting investigation into apolipoproteins (APOs). In this study, we focused on apolipoprotein D (APOD) following comprehensive analyses of APOs in pan-cancer. Utilizing data from the TCGA-STAD and GSE62254 cohorts, we elucidated associations between APOD expression and multiple facets of GC, including prognosis, tumor microenvironment (TME), cancer biomarkers, mutations, and immunotherapy response, and identified potential anti-GC drugs. Single-cell analyses and immunohistochemical staining confirmed APOD expression in fibroblasts within the GC microenvironment. Additionally, we independently validated the prognostic significance of APOD in the ZN-GC cohort. Our comprehensive analyses revealed that high APOD expression in GC patients was notably associated with unfavorable clinical outcomes, reduced microsatellite instability and tumor mutation burden, alterations in the TME, and diminished response to immunotherapy. These findings provide valuable insights into the potential prognostic and therapeutic implications of APOD in GC.

Causal Relationship between Immune Cells and Gynecological Cancers through Bidirectional and Multivariable Mendelian Randomization Analyses.

Background: Evidence suggests potential associations between gynecological malignancies and various immune cell chemicals and systems. However, establishing a causal relationship remains uncertain. Methods: This work employed Wald ratio for one single-nucleotide polymorphism (SNP) or the inverse-variance weighted method (IVW) for multiple SNPs to conduct bidirectional two-sample Mendelian randomization (MR) analysis by utilizing genome-wide association study (GWAS) data. We employed supplementary methods, including MR-Egger and weighted median methods, to detect and correct for the influence of horizontal pleiotropy. In addition, we also use colocalization analysis for further validation. Results: In IVW analysis, increases in relative count of circulating CD11c+ HLA-DR++ conventional dendritic cells (cDC) were associated with an elevated risk of breast cancer (OR [95% CI], 1.1295 [1.0632-1.2000], P = 8.044 × 10-5), while elevated levels of HLA-DR on plasmacytoid dendritic cells (DC) and HLA-DR on DC were protective against breast cancer. In addition, actual count of CD39+ resting Treg AC was also shown to be causally associated with the development of ovarian cancer, whereas a high relative count of CD28+ CD45RA- CD8+ T cells reduced the risk of cervical cancer. Sensitivity analysis revealed almost no evidence of bias in the current study. Multivariable MR (MVMR) analyses further confirmed a direct impact of the CD11c+ HLA-DR++ cDC immune phenotype on breast cancer. Colocalization analysis showed the lead SNP, rs780094, suggesting HLA-DR GWAS shared a common genetic mechanism with breast cancer. Conclusions: The MR study identified significant causal relationships between multiple immunophenotypes and breast cancer, aiming to provide clinicians with some reference for cancer prediction and explore further potential associations between immune phenotypes and gynecologic tumors.

Potential Causal Association between Plasma Metabolites, Immunophenotypes, and Female Reproductive Disorders: A Two-Sample Mendelian Randomization Analysis.

BACKGROUND: While extensive research highlighted the involvement of metabolism and immune cells in female reproductive diseases, causality remains unestablished. METHODS: Instrumental variables for 486 circulating metabolites (N = 7824) and 731 immunophenotypes (N = 3757) were derived from a genome-wide association study (GWAS) meta-analysis. FinnGen contributed data on 14 female reproductive disorders. A bidirectional two-sample Mendelian randomization study was performed to determine the relationships between exposures and outcomes. The robustness of results, potential heterogeneity, and horizontal pleiotropy were examined through sensitivity analysis. RESULTS: High levels of mannose were found to be causally associated with increased risks of gestational diabetes (GDM) (OR [95% CI], 6.02 [2.85-12.73], p = 2.55 × 10-6). A genetically predicted elevation in the relative count of circulating CD28-CD25++CD8+ T cells was causally related to increased female infertility risk (OR [95% CI], 1.26 [1.14-1.40], p = 1.07 × 10-5), whereas a high absolute count of NKT cells reduced the risk of ectopic pregnancy (OR [95% CI], 0.87 [0.82-0.93], p = 5.94 × 10-6). These results remained consistent in sensitivity analyses. CONCLUSIONS: Our study supports mannose as a promising GDM biomarker and intervention target by integrating metabolomics and genomics.

Structurally manipulated antioxidant peptides derived from wheat bran: Preparation and identification.

Bioactive peptide's development is facing two challenges in terms of its lower yield and limited understanding of structurally orientated functionality. Therefore, peptides were prepared from wheat bran via a cocktail enzyme for achieving a higher level of hydrophobic amino acids than traditional method. The obtained peptides exhibited great antioxidant activities against H2O2-induced oxidative stress in HepG2 cells. Among them, 91 bioactive peptides were selected through the virtual screening, and their N-terminal and C-terminal contained many hydrophobic amino acids. Then the peptides with capacity to interact with Keap1 were identified by in silico simulation, because Keap1 acts as a sensor of redox insults. The results revealed that peptides DLDW and DLGL demonstrated the highest binding affinities, and a bridge was formed between Asp of DLGL and Arg415 of Klech domain, contributing to interfering Keap1-Nrf2 interaction. These findings implied a potential application of wheat bran peptides as nutraceuticals and health-promoting ingredients.

Immuno-Enhancing Effect of Ginsenoside Rh2 Liposomes on Foot-and-Mouth Disease Vaccine.

The adjuvant is essential for vaccines because it can enhance or directly induce a strong immune response associated with vaccine antigens. Ginsenoside Rh2 (Rh2) had immunomodulatory effects but was limited by poor solubility and hemolysis. In this study, Rh2 liposomes (Rh2-L) were prepared by ethanol injection methods. The Rh2-L effectively dispersed in a double emulsion adjuvant system to form a Water-in-Oil-in-Water (W/O/W) emulsion and had no hemolysis. The physicochemical properties of the adjuvants were tested, and the immune activity and auxiliary effects indicated by the Foot-and-Mouth disease (FMDV) antigen were evaluated. Compared with the mice vaccinated with the FMD vaccine prepared with the double emulsion adjuvant alone, those with the FMD vaccine prepared with the double emulsion adjuvant containing Rh2-L had significantly higher neutralizing antibody titer and splenocyte proliferation rates and showed higher cellular and humoral immune responses. The results demonstrated that Rh2-L could further enhance the immune effect of the double emulsion adjuvant against Foot-and-Mouth Disease.

Transdermal Microneedles Alleviated Rheumatoid Arthritis by Inducing Immune Tolerance via Skin-Resident Antigen Presenting Cells.

Restoring immune tolerance is the ultimate goal for rheumatoid arthritis (RA) treatment. The most reported oral or intravenous injection routes for the immunization of autoantigens cause gastrointestinal side effects, low patient compliance, and unsatisfied immune tolerance induction. Herein, the use of a transdermal microneedle patch is for the first time investigated to codeliver CII peptide autoantigen and rapamycin for reversing immune disorders of RA. The immunized microneedles efficiently recruit antigen-presenting cells particularly Langerhans cells, and induce tolerogenic dendritic cells at the administration skin site. The tolerogenic dendritic cells further homing to lymph nodes to activate systemic Treg cell differentiation, which upregulates the expression of anti-inflammatory mediators while inhibiting the polarization of Th1/2 and Th17 T cell phenotypes and the expression of inflammatory profiles. As a result, the optimized microneedles nearly completely eliminate RA symptoms and inflammatory infiltrations. Furthermore, it is demonstrated that a low dose of rapamycin is crucial for the successful induction of immune tolerance. The results indicate that a rationally designed microneedle patch is a promising strategy for immune balance restoration with increased immune tolerance induction efficiency and patient compliance.

Structural property of extractable proteins and polysaccharides in wheat bran following a dual-enzymatic pretreatment and corresponding functionality.

The current study applied dual-enzymatic treatment via alcalase and Bacillus velezensis hydrolase for enhancing extraction of proteins and polysaccharides from wheat bran and modifying their corresponding structure. Results indicated the aqueous extract by enzymatic pretreatment (referred as EHWB) had an increased content of soluble substance, in which 18.5 % increased for carbohydrates and 11.4 % increased for proteins in the extract compared to the aqueous extract without enzymes (labeled as AEWB). Furthermore, compositions with lower molecular weight of 130 kDa and < 21.1 kDa for polysaccharides and proteins, respectively, were found in EHWB. Interestingly, EHWB had a twice higher radicals scavenging than that of AEWB, and digestive property indicated EHWB had a greater peptides production although glucose release was lower in gastric phase. Importantly, this is the first study to reveal that gut microbiota fermentation of EHWB resulted in faster generation of short-chain fatty acids at initial fermentation stage (6 h), followed a higher generation of butyrate at final fermentation stage (24 h). This fermentation property might be associated with its presence of lower molecular weight substrates and even the changes in the molecular structure induced by the enzymes. This study highlights a novel approach for developing a value-added product from wheat bran.

An "AND" Logic-Gated Prodrug Micelle Locally Stimulates Antitumor Immunity.

Materials that can respond to multiple biomarkers simultaneously, acting as an "AND" gate, have the potential to enhance tumor-targeting for drug delivery. In this study, an "AND" logic-controlled release prodrug micelle is developed for codelivering the chemotherapeutic and the stimulator of interferon genes (STING) agonist, enabling precise combinatorial therapy. The drug release is programmed by tumor-enriched boramino acids (BAA) in the tumor microenvironment and intracellular reactive oxygen species (ROS), resulting in enhanced tumor targeting. STING agonist is successfully encapsulated into prodrug micelles through π-π stacking and hydrophobic interactions. These AND logic-gated prodrug micelles can achieve tumor-targeted delivery of STING agonist, leading to significantly enhanced immune activation and antitumor efficacy in vivo. It is expected that this clinically relevant nanoplatform will provide a rational design of an effective immunotherapy combination regimen to convert immunologically "cold" tumors to immunogenic "hot" tumors, addressing the major challenges faced by immunotherapies.

Parallel processes of temporal control in the supplementary motor area and the frontoparietal circuit.

Durations in the several seconds' range are cognitively accessible during active timing. Functional neuroimaging studies suggest the engagement of the basal ganglia (BG) and supplementary motor area (SMA). However, their functional relevance and arrangement remain unclear because non-timing cognitive processes temporally coincide with the active timing. To examine the potential contamination by parallel processes, we introduced a sensory control and a motor control to the duration-reproduction task. By comparing their hemodynamic functions, we decomposed the neural activities in multiple brain loci linked to different cognitive processes. Our results show a dissociation of two cortical neural circuits: the SMA for both active timing and motor preparation, followed by a prefrontal-parietal circuit related to duration working memory. We argue that these cortical processes represent duration as the content but at different levels of abstraction, while the subcortical structures, including the BG and thalamus, provide the logistic basis of timing by coordinating the temporal framework across brain structures.

Gut microbiota, host lipid metabolism and regulation mechanism of high-fat diet induced mice following different probiotics-fermented wheat bran intervention.

Wheat bran (WB) was fermented by Lactobacillus rhamnosus, Lactobacillus plantarum, Lactobacillus brevis (LAB-FWB), respectively, and their corresponding mechanism of obesity alleviation via gut microbiota and lipid metabolism was investigated. Results indicated LAB-FWB reduced body weight and serum glucose, followed by an improved lipid profile in obese mice compared with WB. All LAB-FWB interventions led to an enriched steroid hormone biosynthesis. LGG-WB significantly up-regulated genes in arachidonic acid metabolism, bile secretion and linoleic acid metabolism. While LB-WB down-regulated genes in PPAR signaling pathway and LP-WB up-regulated genes in linoleic acid metabolism, indicate their different regulation patterns. Furthermore, LAB-FWB reduced Firmicutes/Bacteroidetes ratio and returned HFD-dependent bacteria Colidextribacter and Erysipelatoclostridium to be normalized. Interestingly, LAB-FWB significantly enriched lipid-related pathways, benefiting xanthohumol, prostaglandin F2alpha, LPI 18:2 and lipoamide biosynthesis in lipid metabolic pathway, but not found in WB group. Among them, treatment with LGG-WB exerted the greatest function on alleviating obesity syndromes.

Pan-cancer analysis revealing that PTPN2 is an indicator of risk stratification for acute myeloid leukemia.

The non-receptor protein tyrosine phosphatases gene family (PTPNs) is involved in the tumorigenesis and development of many cancers, but the role of PTPNs in acute myeloid leukemia (AML) remains unclear. After a comprehensive evaluation on the expression patterns and immunological effects of PTPNs using a pan-cancer analysis based on RNA sequencing data obtained from The Cancer Genome Atlas, the most valuable gene PTPN2 was discovered. Further investigation of the expression patterns of PTPN2 in different tissues and cells showed a robust correlation with AML. PTPN2 was then systematically correlated with immunological signatures in the AML tumor microenvironment and its differential expression was verified using clinical samples. In addition, a prediction model, being validated and compared with other models, was developed in our research. The systematic analysis of PTPN family reveals that the effect of PTPNs on cancer may be correlated to mediating cell cycle-related pathways. It was then found that PTPN2 was highly expressed in hematologic diseases and bone marrow tissues, and its differential expression in AML patients and normal humans was verified by clinical samples. Based on its correlation with immune infiltrates, immunomodulators, and immune checkpoint, PTPN2 was found to be a reliable biomarker in the immunotherapy cohort and a prognostic predictor of AML. And PTPN2'riskscore can accurately predict the prognosis and response of cancer immunotherapy. These findings revealed the correlation between PTPNs and immunophenotype, which may be related to cell cycle. PTPN2 was differentially expressed between clinical AML patients and normal people. It is a diagnostic biomarker and potentially therapeutic target, providing targeted guidance for clinical treatment.

Development of Patient-Reported Outcome Scale for Patients with Diabetic Foot and Its Reliability and Validity Test.

Objective: To construct a self-reported outcome scale for diabetic foot patients, and to test its reliability and validity. Methods: Through literature reading and interviews with 30 patients, a pool of scale items was formed. The items were classified and sorted out according to the expected scale structure framework. After two rounds of expert consultation and a small range of test dressing, the initial scale was formed. Through the investigation of 85 patients with diabetic foot, item differentiation analysis, correlation analysis and exploratory factor analysis were used to screen the items. Cronbach's α coefficient, retest reliability and content and structure validity analysis were used to determine the feasibility and validity of the scale. Results: The final scale included 4 first-level items and 22 second-level items. The critical ratio method showed that the scores of each item in the high group and the low group were significantly different (P < 0.05). Correlation analysis showed that the correlation coefficient between each item and the total score was 0.431 to 0.829; The content validity index of the scale was 0.91, the exploratory factor analysis identified three common factors, and the cumulative variance contribution rate was 75.381%. The confirmatory factor analysis showed that the model fit well. The Cronbach's α coefficient of the scale was 0.934 and the retest reliability coefficient was 0.926. Conclusion: The self-reported outcome scale for diabetic foot patients has good reliability and validity, and can be used to investigate the health status of diabetic foot patients and evaluate the therapeutic effect.

In-modified Sn-MOFs with high catalytic performance in formate electrosynthesis from aqueous carbon dioxide.

Electrochemical reduction of carbon dioxide (CO2ER) has become an effective solution to relieve the energy crisis and tackle climate change. In this study, a series of tin-based organic frameworks modified by In (Sn-MOF/Inx) were successfully synthesized via a simple hydrothermal method and explored for high formate-selective CO2ER. The pure Sn-MOF exhibits maximum formate selectivity with a faradaic efficiency (FEformate) of approximately 85.0% and a current density of 15 mA cm-2 at -1.16 VRHE, while the In (6%)-modified Sn-MOF (Sn-MOF/In6) delivers a much higher maximum FEformate (around 97.5%) and a current density of 16 mA cm-2 at -0.96 VRHE. Remarkably, the Sn-MOF/In6 exhibits a significantly larger specific surface area (183.3 m2 g-1) compared to the Sn-MOF (65.2 m2 g-1). These findings indicate that introducing In, an alien element with a slightly different outer orbital electron number from that of Sn, can significantly boost the selectivity and activity for CO2ER to formate. This study presents an efficient way to modify MOF catalysts through a well-designed introducing process.

Cordycepin inhibits myogenesis via activating the ERK1/2 MAPK signalling pathway in C2C12 cells.

Cordycepin (with a molecular formula of C10H13N5O3), a natural adenosine isolated from Cordyceps militaris, has an important regulatory effect on skeletal muscle remodelling and quality maintenance. The aim of this study was to investigate the effect of cordycepin on myoblast differentiation and explore the underlying molecular mechanisms of this effect. Our results showed that cordycepin inhibited myogenesis by downregulating myogenic differentiation (MyoD) and myogenin (MyoG), preserved undifferentiated reserve cell pools by upregulating myogenic factor 5 (Myf5) and retinoblastoma-like protein p130 (p130), and enhanced energy reserves by decreasing intracellular reactive oxygen species (ROS) and enhancing mitochondrial membrane potential, mitochondrial mass, and ATP content. The effect of cordycepin on myogenesis was associated with increased phosphorylation of extracellular signal-regulated kinase 1/2 (p-ERK1/2). PD98059 (a specific inhibitor of p-ERK1/2) attenuated the inhibitory effect of cordycepin on C2C12 differentiation. The present study reveals that cordycepin inhibits myogenesis through ERK1/2 MAPK signalling activation accompanied by an increase in skeletal muscle energy reserves and improving skeletal muscle oxidative stress, which may have implications for its further application for the prevention and treatment of degenerative muscle diseases caused by the depletion of depleted muscle stem cells.

Chemical characterization, antioxidant and immunomodulatory activities of acetylated polysaccharides from Cyperus esculentus.

This research was designed to characterize the structure of Cyperus esculentus polysaccharide (CEP) and its acetylated one (ACEP), and then investigated the effects of acetylation on the changes in physicochemical properties, thermal stability, antioxidant and immunomodulatory activities. Results showed that CEP and ACEP were heteropolysaccharides consisting of glucose, mannose, arabinose and xylose. The main chain of CEP included α-1,4-Glcp residues with the branching points at the O-6 position of the α-1,6-Manp residues. Acetyl groups were substituted at the O-2 and O-6 positions of some glucose residues. Meanwhile, the acetylation remarkably improved the polysaccharides thermal stability, and the ACEP exhibited a greater antioxidant activity. Furthermore, CEP and ACEP were proved to protect RAW 264.7 cells against LPS-induced inflammation by improving cellular morphology and decreasing reactive oxygen species secretion. This study may highlight a new approach for developing a high value-added ingredient from C. esculentus for functional food industry.

Impact of de-branched starch molecules and fatty acid complexes on the attenuation of ageing-induced cognitive impairment.

BACKGROUND: Ageing and associated cognitive impairments are becoming serious issues around the world. In this study, the physiological properties of three kinds of complexes of fatty acid (capric, stearic and oleic acid, respectively) and de-branched starch molecules were investigated via a d-galactose-induced ageing model. This study revealed differences in the regulation of cognitive impairment and brain damage following intervention of different complexes, which might highlight a potent approach for the prevention of this chronic disease. RESULTS: Data indicated that three complexes improved response time and cognitive function and the bio-parameter markers associated with oxidative stress in ageing rats. Among them, the complexes prepared from de-branched starch-oleic acid showed a greater improvement compared to others. In addition, de-branched starch-capric acid complex showed a higher improvement in the morphology of colon cells and hippocampal neuronal cells. The consumption of de-branched starch-capric acid and -oleic acid complexes generated more short-chain fatty acids in the gut. More importantly, the complexation of de-branched starch with either caprate or stearate enhanced gut Akkermansia. Therefore, it was proposed that the richness in Akkermansia and gut metabolites might be associated with reduced damage of the hippocampal neuronal cells induced by the ageing progress. Moreover, the AMPK (AMP-activated protein kinase) pathway was activated in liver in de-branched starch-capric acid complex diet. In summary, de-branched starch-capric acid complex exhibited a greater effect on the attenuation of ageing-induced cognitive impairment. CONCLUSION: This study might highlight a new approach for intervening in the cognitive impairment during the ageing progress via a food supply. © 2023 Society of Chemical Industry.

Periodic Acid Modification of Chemical-Bath Deposited SnO2 Electron Transport Layers for Perovskite Solar Cells and Mini Modules.

Chemical bath deposition (CBD) has been demonstrated as a remarkable technology to fabricate high-quality SnO2 electron transport layer (ETL) for large-area perovskite solar cells (PSCs). However, surface defects always exist on the SnO2 film coated by the CBD process, impairing the devices' performance. Here, a facile periodic acid post-treatment (PAPT) method is developed to modify the SnO2 layer. Periodic acid can react with hydroxyl groups on the surface of SnO2 films and oxidize Tin(II) oxide to Tin(IV) oxide. With the help of periodic acid, a better energy level alignment between the SnO2 and perovskite layers is achieved. In addition, the PAPT method inhibits interfacial nonradiative recombination and facilitates charge transportation. Such a multifunctional strategy enables to fabricate PSC with a champion power conversion efficiency (PCE) of 22.25%, which remains 93.32% of its initial efficiency after 3000 h without any encapsulation. Furthermore, 3 × 3 cm2 perovskite mini-modules are presented, achieving a champion efficiency of 18.10%. All these results suggest that the PAPT method is promising for promoting the commercial application of large-area PSCs.