A Monte Carlo simulation-based health risk assessment of heavy metals in soils of the tropical region in southern China.

The disturbance of ecological stability may take place in tropical regions due to the elevated biomass density resulting from heavy metal and other contaminant pollution. In this study, 62 valid soil samples were collected from Sanya. Source analysis of heavy metals in the area was carried out using absolute principal component-multiple linear regression receptor modelling (APCS-MLR); the comprehensive ecological risk of the study area was assessed based on pollution sources; the Monte-Carlo model was used to accurately predict the health risk of pollution sources in the study area. The results showed that: The average contents of soil heavy metals Cu, Ni and Cd in Sanya were 5.53, 6.56 and 11.66 times higher than the background values of heavy metals. The results of soil geo-accumulation index (Igeo) showed that Cr, Mo, Mn and Zn were unpolluted to moderately polluted, Cu and Ni were moderately polluted, and Cd was moderately polluted to strongly polluted. The main sources of heavy metal pollution were natural sources (57.99%), agricultural sources (38.44%) and traffic sources (3.57%). Natural and agricultural sources were jointly identified as priority control pollution sources and Cd was the priority control pollution element for soil ecological risk. Heavy metal content in Sanya did not pose a non-carcinogenic risk to the population, but there was a carcinogenic risk to children. The element Zn had a high carcinogenic risk to children, and was a priority controlling pollutant element for the risk of human health, with agricultural sources as the priority controlling pollutant source.

Effects of blood-processing protocols on cell-free DNA fragmentomics in plasma: Comparisons of one- and two-step centrifugations.

BACKGROUND: Cell-free DNA (cfDNA) fragmentomic characteristics are promising analytes with abundant physiological signals for non-invasive disease diagnosis and monitoring. Previous studies on plasma cfDNA fragmentomics commonly employed a two-step centrifugation process for removing cell debris, involving a low-speed centrifugation followed by a high-speed centrifugation. However, the effects of centrifugation conditions on the analysis of cfDNA fragmentome remain uncertain. METHODS: We collected blood samples from 10 healthy individuals and divided each sample into two aliquots for plasma preparation with one- and two-step centrifugation processes. We performed whole genome sequencing (WGS) of the plasma cfDNA in the two groups and comprehensively compared the cfDNA fragmentomic features. Additionally, we reanalyzed the fragmentomic features of cfDNA from 16 healthy individuals and 16 COVID-19 patients, processed through one- and two-step centrifugation in our previous study, to investigate the impact of centrifugation on disease signals. RESULTS: Our results showed that there were no significant differences observed in the characteristics of nuclear cfDNA, including size, motif diversity score (MDS) of end motifs, and genome distribution, between plasma samples treated with one- and two-step centrifugation. The cfDNA size shortening in COVID-19 patients was observed in plasma samples with one- and two-step centrifugation methods. However, we observed a significantly higher relative abundance and longer size of cell-free mitochondrial DNA (mtDNA) in the one-step samples compared to the two-step samples. This difference in mtDNA caused by the one- and two-step centrifugation methods surpasses the pathological difference between COVID-19 patients and healthy individuals. CONCLUSIONS: Our findings indicate that one-step low-speed centrifugation is a simple and potentially suitable method for analyzing nuclear cfDNA fragmentation characteristics. These results offer valuable guidance for cfDNA research in various clinical scenarios.

Prolonged Activity Deprivation Causes Pre- and Postsynaptic Compensatory Plasticity at Neocortical Excitatory Synapses.

Homeostatic plasticity stabilizes firing rates of neurons, but the pressure to restore low activity rates can significantly alter synaptic and cellular properties. Most previous studies of homeostatic readjustment to complete activity silencing in rodent forebrain have examined changes after 2 d of deprivation, but it is known that longer periods of deprivation can produce adverse effects. To better understand the mechanisms underlying these effects and to address how presynaptic as well as postsynaptic compartments change during homeostatic plasticity, we subjected mouse cortical slice cultures to a more severe 5 d deprivation paradigm. We developed and validated a computational framework to measure the number and morphology of presynaptic and postsynaptic compartments from super-resolution light microscopy images of dense cortical tissue. Using these tools, combined with electrophysiological miniature excitatory postsynaptic current measurements, and synaptic imaging at the electron microscopy level, we assessed the functional and morphological results of prolonged deprivation. Excitatory synapses were strengthened both presynaptically and postsynaptically. Surprisingly, we also observed a decrement in the density of excitatory synapses, both as measured from colocalized staining of pre- and postsynaptic proteins in tissue and from the number of dendritic spines. Overall, our results suggest that cortical networks deprived of activity progressively move toward a smaller population of stronger synapses.

circRNA6448-14/miR-455-3p/OTUB2 axis stimulates glycolysis and stemness of esophageal squamous cell carcinoma.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) is a gastrointestinal malignancy with high incidence. This study aimed to reveal the complete circRNA-miRNA-mRNA regulatory network in ESCC and validate its function mechanism. METHOD: Expression of OTU Domain-Containing Ubiquitin Aldehyde-Binding Protein 2 (OTUB2) in ESCC was analyzed by bioinformatics to find the binding sites between circRNA6448-14 and miR-455-3p, as well as miR-455-3p and OTUB2. The binding relationships were verified by RNA Immunoprecipitation (RIP) and dual-luciferase assay. The expressions of circRNA6448-14, miR-455-3p, and OTUB2 were detected by quantitative real-time polymerase chain reaction (qRT-PCR). MTT assay measured cell viability, and the spheroid formation assay assessed the ability of stem cell sphere formation. Western blot (WB) determined the expression of marker proteins of stem cell surface and rate-limiting enzyme of glycolysis. The Seahorse XFe96 extracellular flux analyzer measured the rate of extracellular acidification rate and cellular oxygen consumption. Corresponding assay kits assessed cellular glucose consumption, lactate production, and adenosine triphosphate (ATP) generation. RESULTS: In ESCC, circRNA6448-14 and OTUB2 were highly expressed in contrast to miR-455-3p. Knocking down circRNA6448-14 could prevent the glycolysis and stemness of ESCC cells. Additionally, circRNA6448-14 enhanced the expression of OTUB2 by sponging miR-455-3p. Overexpression of OTUB2 or silencing miR-455-3p reversed the inhibitory effect of knockdown of circRNA6448-14 on ESCC glycolysis and stemness. CONCLUSION: This research demonstrated that the circRNA6448-14/miR-455-3p/OTUB2 axis induced the glycolysis and stemness of ESCC cells. Our study revealed a novel function of circRNA6448-14, which may serve as a potential therapeutic target for ESCC.

Efficacy and safety of Zhishixiaopi decoction in functional dyspepsia: A meta-analysis of randomized controlled trials.

BACKGROUND: Functional dyspepsia (FD) refers to a group of clinical symptoms caused by gastric and duodenal dysfunction. Which is a chronic functional disorder of the gastrointestinal tract with no cure. Zhishixiaopi decoction (ZSXP) is a type of Chinese herbal prescription that for treating FD. Although some randomized controlled trials (RCTs) report that ZSXP can significantly improve FD clinical symptoms and/or laboratory results, the trial design varies greatly among studies, making it challenging to draw a conclusion of the efficacy of ZSXP in treating FD. DESIGN: A systematic review and a meta-analysis. SETTING: Mianyang Central Hospital. OBJECTIVE: We conducted a systematic review and a meta-analysis to evaluate the efficacy and safety of ZSXP for treating FD. METHODS: We developed inclusion and exclusion criteria based on FD diagnosed criteria, interventions to treat FD, and outcomes of these interventions. Search strategies combined disease terms, symptom terms, anatomy terms and intervention terms. Literature search was conducted on eight online databases in English or Chinese, including Medline (via PubMed), Embase (via Ovid), The Cochrane Library, Web of Science, China Biology Medicine (CBM), China National Knowledge Infrastructure (CNKI), Chinese Scientific Journals Database (VIP), and Wanfang Database. INTERVENTION: The experimental group received oral administration of ZSXP and had a complete treatment process. ZSXP needs to fully contain the key herbal ingredients, regardless of whether the dosage of each herb is consistent with the original prescription. The Control group received monotherapy or combination therapy of other Western medicine and had a complete treatment process. OUTCOMES: The primary outcomes appraised were Total effective rate (TER), serum levels of Motilin(MOT), Gastrin(GAS) and Somatostatin (SS), Gastric emptying rate (GER) using a Barium meal method (GER(B)) and Gastric half emptying time using an Ultrasonic method (GHET(T1/2)). The Cochrane Bias Risk Tool was used for quality critical appraisal, Review Manager (RevMan) version 5.3 was used for statistical analysis. RESULTS: A total of 21 medium-quality RCTs were included in the meta-analysis. All 21 included studies were conducted and completed in Mainland China from 1998 to 2020. The treatment duration was between two weeks to two months. The meta-analysis suggests that, compared with the Western medicine treatment group, ZSXP treatment was more effective to improving the TER in FD [Odds ratio, OR = 3.54, 95%CI:(2.49, 5.05), Z = 6.99, P<0.00001] without significant increase in adverse events. However, no statistical significance was found between the groups in serum MOT levels [Standard mean difference, SMD = 1.05, 95%CI:(-0.42, 2.53), Z = 1.04, P = 0.16], serum GAS levels [SMD = -0.16, 95%CI:(-1.20, 0.88), Z = 0.31, P = 0.76], serum SS levels [SMD = -0.04, 95%CI:(-1.97, 1.89), Z = 0.04, P = 0.97], GER(B) [SMD = 1.09, 95%CI:(-0.81, 3.00), Z = 1.12, P = 0.26]or GHET(T1/2) [Mean difference, MD = -2.18, 95%CI:(-5.55, 1.19), Z = 1.27, P = 0.20]. CONCLUSIONS: The meta-analysis suggests that Zhishixiaopi treatment is a relatively effective and safe traditional Chinese medicine prescription and could be used for functional dyspepsia treatment. Considering the limitations of this study, the conclusion needs to be further confirmed by high-quality, multi-center, and large-sample randomized controlled trials.

Cryo-EM structure of human HCN3 channel and its regulation by cAMP.

HCN channels are important for regulating heart rhythm and nerve activity and have been studied as potential drug targets for treating depression, arrhythmia, nerve pain, and epilepsy. Despite possessing unique pharmacological properties, HCN channels share common characteristics in that they are activated by hyperpolarization and modulated by cAMP and other membrane lipids. However, the mechanisms of how these ligands bind and modulate HCN channels are unclear. In this study, we solved structures of full-length human HCN3 using cryo-EM and captured two different states, including a state without any ligand bound and a state with cAMP bound. Our structures reveal the novel binding sites for cholesteryl hemisuccinate in apo state and show how cholesteryl hemisuccinate and cAMP binding cause conformational changes in different states. These findings explain how these small modulators are sensed in mammals at the molecular level. The results of our study could help to design more potent and specific compounds to influence HCN channel activity and offer new therapeutic possibilities for diseases that lack effective treatment.

Mechanical biomimetic silk nano fiber-magnesium ion complex/hydroxyethylcellulose/glycerol hydrogel dressing with angiogenic capacity for accelerating scarless diabetic wound healing.

Quick scarless healing remains a key issue for diabetic wounds. Here, a stretchable elastomeric hydrogel dressing composed of hydroxyethylcellulose (HEC), silk nano fiber-magnesium ion complex (Mg2+-SNF) and glycerol (Gly) was developed to optimize mechanical niche, anti-inflammatory and angiogenic behavior simultaneously. The composite hydrogel dressing exhibited skin-like elasticity (175.1 ± 23.9 %) and modulus (156.7 ± 2.5 KPa) while Mg2+-SNF complex endowed the dressing with angiogenesis, both favoring quick scarless skin regeneration. In vitro cell studies revealed that the hydrogel dressing stimulated fibroblast proliferation, endothelial cell migration and vessel-like tube formation, and also induced anti-inflammatory behavior of macrophages. In vivo results revealed accelerated healing of diabetic wounds. The improved granulation ingrowth and collagen deposition suggested high quality repair. Both thinner epidermal layer and low collagen I/III ratio of the regenerated skin confirmed scarless tissue formation. This bioactive hydrogel dressing has promising potential to address the multifaceted challenges of diabetic wound management.

Prediction of Junior High School Students' Problematic Internet Use: The Comparison of Neural Network Models and Linear Mixed Models in Longitudinal Study.

Purpose: With the rise of big data, deep learning neural networks have garnered attention from psychology researchers due to their ability to process vast amounts of data and achieve superior model fitting. We aim to explore the predictive accuracy of neural network models and linear mixed models in tracking data when subjective variables are predominant in the field of psychology. We separately analyzed the predictive accuracy of both models and conduct a comparative study to further investigate. Simultaneously, we utilized the neural network model to examine the influencing factors of problematic internet usage and its temporal changes, attempting to provide insights for early interventions in problematic internet use. Patients and Methods: This study compared longitudinal data of junior high school students using both a linear mixed model and a neural network model to ascertain the efficacy of these two methods in processing psychological longitudinal data. Results: The neural network model exhibited significantly smaller errors compared to the linear mixed model. Furthermore, the outcomes from the neural network model revealed that, when analyzing data from a single time point, the influences of seventh grade better predicted Problematic Internet Use in ninth grade. And when analyzing data from multiple time points, the influences of sixth, seventh, and eighth grades more accurately predicted Problematic Internet Use in ninth grade. Conclusion: Neural network models surpass linear mixed models in precision when predicting and analyzing longitudinal data. Furthermore, the influencing factors in lower grades provide more accurate predictions of Problematic Internet Use in higher grades. The highest prediction accuracy is attained through the utilization of data from multiple time points.

Total syntheses of Tetrodotoxin and 9-epiTetrodotoxin.

Tetrodotoxin and congeners are specific voltage-gated sodium channel blockers that exhibit remarkable anesthetic and analgesic effects. Here, we present a scalable asymmetric syntheses of Tetrodotoxin and 9-epiTetrodotoxin from the abundant chemical feedstock furfuryl alcohol. The optically pure cyclohexane skeleton is assembled via a stereoselective Diels-Alder reaction. The dense heteroatom substituents are established sequentially by a series of functional group interconversions on highly oxygenated cyclohexane frameworks, including a chemoselective cyclic anhydride opening, and a decarboxylative hydroxylation. An innovative SmI2-mediated concurrent fragmentation, an oxo-bridge ring opening and ester reduction followed by an Upjohn dihydroxylation deliver the highly oxidized skeleton. Ruthenium-catalyzed oxidative alkyne cleavage and formation of the hemiaminal and orthoester under acidic conditions enable the rapid assembly of Tetrodotoxin, anhydro-Tetrodotoxin, 9-epiTetrodotoxin, and 9-epi lactone-Tetrodotoxin.

The value of color Doppler ultrasonography combined with serum tumor markers in differential diagnosis of gastric stromal tumor and gastric cancer.

This study aimed to explore the value of color Doppler ultrasonography combined with carcinoembryonic antigen (CEA) and carbohydrate antigen 19-9 (CA19-9) in differential diagnosis of gastric stromal tumor (GST) and gastric cancer (GC). An analysis of the clinical data of 180 patients with clinically suspected gastric space occupying lesions. According to the postoperative pathological results, 180 suspected gastric space-occupying lesion patients were divided into GST group (n = 83) and GC group (n = 97). Color Doppler ultrasonography, serum tumor markers CEA and CA19-9 were compared. The research results showed that serum CEA and CA19-9 levels were lower in patients with GST group than those with GC group (both P < 0.001). With postoperative pathology as the gold standard, detection rates of GST and GC by combination of color Doppler ultrasound (CDUS), serum CEA, and CA19-9 were higher than those of each index alone (both P < 0.001). There was no difference between detection rates of GST and GC by combination of CDUS, serum CEA, and CA19-9 (P = 0.058). Color Doppler ultrasonography combined with serum tumor markers CEA and CA19-9 tests has a certain differential diagnostic value for GST and GC, which may provide a reliable reference basis for clinical diagnosis and treatment.

[Spatial Distribution of Aerobic Bacteria, Sources and Risks of Nitrogen and Phosphorus in Taihu Lake Sediments].

The release of nitrogen and phosphorus from sediments into lake water will exacerbate the eutrophication of lakes and endanger ecological safety and human health. Microorganisms are indispensable in nitrogen and phosphorus conversion, and accurate analysis of the distribution characteristics and sources of nitrogen and phosphorus in sediments as well as their relationship with microorganisms is an important prerequisite for lake eutrophication control. Taking Taihu Lake as the study area, 30 surface sediment samples were collected, and the grain size, pH, organic matter (OM), dissolved organic carbon (DOC), total phosphorus (TP), total nitrogen (TN), nitrate nitrogen (NO3--N), and dissolved organic nitrogen (DON) along with some other index contents were measured and analyzed; accordingly, spatial distribution characteristics were analyzed. While using nutrient agar (NA), the number of aerobic bacteria (AB) was determined by plate counting in the medium. Combined with principal component analysis (PCA) and Pearson correlation analysis, the spatial distribution characteristics and sources of sediments and AB in Taihu Lake were explored. The characteristics of sediment pollution in Taihu Lake were studied using the comprehensive pollution index and the organic pollution index methods. The results revealed that the average sediment indicators of the surface layer of Taihu Lake were as follows:AB was 9.25×104 CFU·g-1, average particle size (MZ) was 17.59 µm, pH was 7.62, ω(OM) was 15.05 g·kg-1, ω(DOC) was 71.60 mg·kg-1, ω(TP) was 598.13 mg·kg-1, ω(TN) was 1113.92 mg·kg-1, ω(NO3--N) was 3.22 mg·kg-1, and ω(DON) was 22.60 mg·kg-1. The comprehensive pollution index (FF) showed that 13% of the Taihu Lake was moderately polluted, while 87% was heavily polluted. Excluding the area in the center of the lake, the southern lake area, and some lakes in the western part of the East Taihu Lake, TN in the rest of the area was moderately and severely polluted. In addition to the heavy pollution of Zhushan Bay, the TP in Taihu Lake was generally at light and moderate pollution. The organic pollution index (OI) showed that the organic pollution of the sediments of Taihu Lake was relatively light, majorly caused by organic nitrogen (ON) pollution. DOC, DON, TN, and OM in Taihu Lake were primarily derived from the influence of aquatic plants, and TP And AB were primarily derived from the influence of the external input of rivers. This research will provide theoretical support for lake eutrophication treatment and also provide new ideas for further analysis of AB to remove nitrogen and phosphorus pollution from sediments.

Problematic Internet Use in Early Adolescents: Gender and Loneliness Differences in a Latent Growth Model.

Purpose: Based on the Cognitive-Behavioral model and the Interaction of Person-Affect-Cognition-Execution model, this study examined the developmental trajectory of problematic Internet use (PIU) in early adolescents and explored whether there were gender differences in the onset level and rate of development of this developmental trajectory, and tested whether developmental changes in loneliness could have an impact on the developmental trajectory of problematic Internet use. Participants and Methods: This longitudinal study collected data on PIU and loneliness from 296 early adolescents (Mage=11.65, SD=0.58) in four waves. The development of PIU in adolescents and the effects of gender and loneliness development on PIU development were examined using a latent growth model. Results: The results revealed that individuals' PIU development showed a nonlinear latent growth model, with PIU significantly higher than 0 in grade 6 and its growth rate slowing down as PIU increased. Individuals' PIU at low starting levels developed more rapidly later. Boys had higher initial levels of PIU but their PIU developed and increased at the same rate as girls'. Both the initial value and slope of loneliness had a significant effect on the initial value and slope of boys' and girls' PIU. Conclusion: Interventions for PIU in early adolescents also need to consider loneliness at the same time, and intervention groups can focus on individuals with low initial levels of PIU, boys, and individuals with high levels of loneliness.

[Occurrence Characteristics of Microplastics in Baiyangdian Lake Water and Sediments].

In order to explore the occurrence characteristics of microplastics in the freshwater environment of Baiyangdian Lake in China, ten overlying water samples and ten sediment samples were collected in Baiyangdian Lake of Hebei Province in October 2021, and the abundance distribution, shape, particle size, and polymer type of microplastics in the samples were identified using laboratory pretreatment, microscope observation, and laser infrared spectroscopy. The sedimentation law of microplastics at the overlying water-sediment interface was studied using the Stokes sedimentation formula, and their pollution characteristics and potential sources were analyzed. The abundances of microplastics in the overlying water and sediments in Baiyangdian Lake ranged from 474-19382 n·m-3 and 95.3-29542.5 n·kg-1, respectively, with an average value of 6255.4 n·m-3 and 11088 n·kg-1. The main polymer of the microplastics in the overlying water was polyethylene terephthalate[PET, (17.20±0.26)%], and the microplastics in the sediments were mainly chlorinated polyethylene[CPE, (46.11±1.30)%]. The sedimentation velocities of microplastics in the sedimentation zone ranged from 0.0793-111.7547 mm·s-1. The particles with larger particle size had higher sedimentation velocity and easily settled and remained in the sediments. The main sources of microplastic pollution in the study area were the discharge of textile fibers from washing wastewater and the wear and tear of ship paint, ship rubber, and building materials.

MD-SGT: Multi-dilation spherical graph transformer for unsupervised medical image registration.

Deformable medical image registration is an essential preprocess step for several clinical applications. Even though the existing convolutional neural network and transformer based methods achieved the promising results, the limited long-range spatial dependence and non-uniform attention span of these models prohibit further improving the registration performance. To deal with this issue, we proposed a multi-dilation spherical graph transformer (MD-SGT), in which the encoder combined the advantages of convolutional and graph transformer blocks to distinguish effectively the differences between the reference and the template images at various scales. Specifically, the features of each voxel were obtained by aggregating the information from its neighbors sampled from different spherical regions with different dilation rates. The implicit convolution inductive bias and long-range uniform attention span induced by such information aggregation manner made the features more representative for registration. Through the qualitative and quantitative comparisons with state-of-the-art methods on two datasets, we demonstrated that combining long-range uniform attention span and inductive bias are beneficial for promoting the image registration performance, with the Dice score, ASD and HD95 being improved at least by 0.5%, 2.2% and 1.1%, respectively.

Tumor-associated myeloid cells in cancer immunotherapy.

Tumor-associated myeloid cells (TAMCs) are among the most important immune cell populations in the tumor microenvironment, and play a significant role on the efficacy of immune checkpoint blockade. Understanding the origin of TAMCs was found to be the essential to determining their functional heterogeneity and, developing cancer immunotherapy strategies. While myeloid-biased differentiation in the bone marrow has been traditionally considered as the primary source of TAMCs, the abnormal differentiation of splenic hematopoietic stem and progenitor cells, erythroid progenitor cells, and B precursor cells in the spleen, as well as embryo-derived TAMCs, have been depicted as important origins of TAMCs. This review article provides an overview of the literature with a focus on the recent research progress evaluating the heterogeneity of TAMCs origins. Moreover, this review summarizes the major therapeutic strategies targeting TAMCs with heterogeneous sources, shedding light on their implications for cancer antitumor immunotherapies.

Quantitative Proteomics-Based Substrate Screening Revealed Cyclophilin Stabilization Regulated by Deubiquitinase Ubp7.

Quantitative proteomics has emerged as a crucial approach to identifying ubiquitinated substrates to investigate the functions of ubiquitination in cells. In this regard, although the substrate screening of certain enzymes in the ubiquitin system has been based on proteome or ubiquitinome level measurements, the direct comparison of these two approaches has not been determined to date. To quantitatively compare the efficiency and effectiveness of substrate screening from the entire proteomics to the ubiquitinomics filter, we used yeast deubiquitinating enzyme, Ubp7, as an example to evaluate it in this study. A total of 112 potential ubiquitinated substrates were identified from the ubiquitinomics level, whereas only 27 regulated substrates were identified from the entire proteomic screening, demonstrating the increased efficiency of ubiquitinomics quantitative analysis. Subsequently, we selected cyclophilin A (Cpr1) protein as an example, which was filtered out at the proteomics level but was a promising candidate according to the ubiquitinomics filter. Additional investigations revealed that Cpr1 possessed a K48-linked ubiquitin chain regulated by Ubp7, which may affect its homeostasis and, consequently, sensitivity to the therapeutic drug cyclosporine (CsA).

Quantitative proteomics revealed the transition of ergosterol biosynthesis and drug transporters processes during the development of fungal fluconazole resistance.

Fungal infections and antifungal resistance are the increasing global public health concerns. Mechanisms of fungal resistance include alterations in drug-target interactions, detoxification by high expression of drug efflux transporters, and permeability barriers associated with biofilms. However, the systematic panorama and dynamic changes of the relevant biological processes of fungal drug resistance acquisition remain limited. In this study, we developed a yeast model of resistance to prolonged fluconazole treatment and utilized the isobaric labels TMT (tandem mass tag)-based quantitative proteomics to analyze the proteome composition and changes in native, short-time fluconazole stimulated and drug-resistant strains. The proteome exhibited significant dynamic range at the beginning of treatment but returned to normal condition upon acquisition drug resistance. The sterol pathway responded strongly under a short time of fluconazole treatment, with increased transcript levels of most enzymes facilitating greater protein expression. With the drug resistance acquisition, the sterol pathway returned to normal state, while the expression of efflux pump proteins increased obviously on the transcription level. Finally, multiple efflux pump proteins showed high expression in drug-resistant strain. Thus, families of sterol pathway and efflux pump proteins, which are closely associated with drug resistance mechanisms, may play different roles at different nodes in the process of drug resistance acquisition. Our findings uncover the relatively important role of efflux pump proteins in the acquisition of fluconazole resistance and highlight its potential as the vital antifungal targets.

Construction of Probiotic Double-Layered Multinucleated Microcapsules Based on Sulfhydryl-Modified Carboxymethyl Cellulose Sodium for Increased Intestinal Adhesion of Probiotics and Therapy for Intestinal Inflammation Induced by Escherichia coli O157:H7.

The decreased number of viable bacteria and the ability of Bifidobacterium to adhere to and colonize the gut in the gastrointestinal environment greatly limit their efficacy. To solve this problem, thiolated carboxymethyl cellulose sodium (CMC) probiotic double-layered multinucleated microcapsules with Bifidobacterium adolescentis FS2-3 in the inner layer and Bacillus subtilis SN15-2 embedded in the outer layers were designed. First, the viable counts and release rates of microcapsules were examined by in vitro simulated digestion assays, and it was found that microcapsules were better protected from gastrointestinal digestion than the controls. Compared with free Bifidobacterium strains, double-layered multinucleated microcapsules have higher viable bacterial survival rates and storage stability. Second, through in vitro rheology, tensile tests, isotherm titration calorimetry, and adhesion tests, it was observed that thiolated CMC could enhance the strong interaction of Bifidobacterium with intestinal mucus and significantly promote the proliferation and growth of probiotics. Finally, double-layered multinucleated microcapsules containing B. adolescentis FS2-3 and B. subtilis SN15-2 modified with sulfhydryl-modified CMC were studied in the intestine. Alleviation of Escherichia coli O157:H7 induced intestinal inflammation. The results showed that microencapsulation could significantly increase the colon content of Bifidobacterium, relieve intestinal inflammation symptoms in mice with bacterial enteritis, and repair the intestinal microbiota disorder caused by inflammation. The probiotic double-layered multinucleated microcapsules prepared in this study can improve the survival rate of probiotics and promote proliferation, adhesion, and colonization of probiotics.