Early detection of uterine corpus endometrial carcinoma utilizing plasma cfDNA fragmentomics.

BACKGROUND: Uterine corpus endometrial carcinoma (UCEC) is a prevalent gynecologic malignancy with a favorable prognosis if detected early. However, there is a lack of accurate and reliable early detection tests for UCEC. This study aims to develop a precise and non-invasive diagnostic method for UCEC using circulating cell-free DNA (cfDNA) fragmentomics. METHODS: Peripheral blood samples were collected from all participants, and cfDNA was extracted for analysis. Low-coverage whole-genome sequencing was performed to obtain cfDNA fragmentomics data. A robust machine learning model was developed using these features to differentiate between UCEC and healthy conditions. RESULTS: The cfDNA fragmentomics-based model showed high predictive power for UCEC detection in training (n = 133; AUC 0.991) and validation cohorts (n = 89; AUC 0.994). The model manifested a specificity of 95.5% and a sensitivity of 98.5% in the training cohort, and a specificity of 95.5% and a sensitivity of 97.8% in the validation cohort. Physiological variables and preanalytical procedures had no significant impact on the classifier's outcomes. In terms of clinical benefit, our model would identify 99% of Chinese UCEC patients at stage I, compared to 21% under standard care, potentially raising the 5-year survival rate from 84 to 95%. CONCLUSION: This study presents a novel approach for the early detection of UCEC using cfDNA fragmentomics and machine learning showing promising sensitivity and specificity. Using this model in clinical practice could significantly improve UCEC management and control, enabling early intervention and better patient outcomes. Further optimization and validation of this approach are warranted to establish its clinical utility.

Poly(Photosensitizer-Prodrug) Unimolecular Micelles for Chemo-Photodynamic Synergistic Therapy of Antitumor and Antibacteria.

It is crucial to use simple methods to prepare stable polymeric micelles with multiple functions for cancer treatment. Herein, via a "bottom-up" strategy, we reported the fabrication of ß-CD-(PEOSMA-PCPTMA-PPEGMA)21 (ßPECP) unimolecular micelles that could simultaneously treat tumors and bacteria with chemotherapy and photodynamic therapy (PDT). The unimolecular micelles consisted of a 21-arm ß-cyclodextrin (ß-CD) core as a macromolecular initiator, photosensitizer eosin Y (EOS-Y) monomer EOSMA, anticancer drug camptothecin (CPT) monomer, and a hydrophilic shell PEGMA. Camptothecin monomer (CPTMA) could achieve controlled release of the CPT due to the presence of responsively broken disulfide bonds. PEGMA enhanced the biocompatibility of micelles as a hydrophilic shell. Two ßPECP with different lengths were synthesized by modulating reaction conditions and the proportion of monomers, which both were self-assembled to unimolecular micelles in water. ßPECP unimolecular micelles with higher EOS-Y/CPT content exhibited more excellent 1O2 production, in vitro drug release efficiency, higher cytotoxicity, and superior antibacterial activity. Also, we carried out simulations of the self-assembly and CPT release process of micelles, which agreed with the experiments. This nanosystem, which combines antimicrobial and antitumor functions, provides new ideas for bacteria-mediated tumor clinical chemoresistance.

Sesamolin serves as an MYH14 inhibitor to sensitize endometrial cancer to chemotherapy and endocrine therapy via suppressing MYH9/GSK3ß/ß-catenin signaling.

BACKGROUND: Endometrial cancer (EC) is one of the most common gynecological cancers. Herein, we aimed to define the role of specific myosin family members in EC because this protein family is involved in the progression of various cancers. METHODS: Bioinformatics analyses were performed to reveal EC patients' prognosis-associated genes in patients with EC. Furthermore, colony formation, immunofluorescence, cell counting kit 8, wound healing, and transwell assays as well as coimmunoprecipitation, cycloheximide chase, luciferase reporter, and cellular thermal shift assays were performed to functionally and mechanistically analyze human EC samples, cell lines, and a mouse model, respectively. RESULTS: Machine learning techniques identified MYH14, a member of the myosin family, as the prognosis-associated gene in patients with EC. Furthermore, bioinformatics analyses based on public databases showed that MYH14 was associated with EC chemoresistance. Moreover, immunohistochemistry validated MYH14 upregulation in EC cases compared with that in normal controls and confirmed that MYH14 was an independent and unfavorable prognostic indicator of EC. MYH14 impaired cell sensitivity to carboplatin, paclitaxel, and progesterone, and increased cell proliferation and metastasis in EC. The mechanistic study showed that MYH14 interacted with MYH9 and impaired GSK3ß-mediated ß-catenin ubiquitination and degradation, thus facilitating the Wnt/ß-catenin signaling pathway and epithelial-mesenchymal transition. Sesamolin, a natural compound extracted from Sesamum indicum (L.), directly targeted MYH14 and attenuated EC progression. Additionally, the compound disrupted the interplay between MYH14 and MYH9 and repressed MYH9-regulated Wnt/ß-catenin signaling. The in vivo study further verified sesamolin as a therapeutic drug without side effects. CONCLUSIONS: Herein, we identified that EC prognosis-associated MYH14 was independently responsible for poor overall survival time of patients, and it augmented EC progression by activating Wnt/ß-catenin signaling. Targeting MYH14 by sesamolin, a cytotoxicity-based approach, can be applied synergistically with chemotherapy and endocrine therapy to eventually mitigate EC development. This study emphasizes MYH14 as a potential target and sesamolin as a valuable natural drug for EC therapy.

Cadmium transfer between maize and soybean plants via common mycorrhizal networks.

More than 80% terrestrial plants establish mutualistic symbiosis with soil-borne arbuscular mycorrhizal fungi (AMF). These fungi not only significantly improve plant nutrient acquisition and stress resistance, but also mitigate heavy metal phytotoxicity, Furthermore, the extraradical mycorrhizal mycelia can form common mycorrhizal networks (CMNs) that link roots of multiple plants in a community. Here we show that the networks mediate migration of heavy metal cadmium (Cd) from maize (Zea mays L.) to soybean (Glycine max (Linn.) Merr.) plants. CMNs between maize and soybean plants were established after inoculation of maize plants with AMF Funneliformis mosseae. Application of CdCl2 in maize plants led to 64.4% increase in the shoots and 48.2% increase in the roots in Cd content in CMNs-connected soybean plants compared to the control without Cd treatment in maize. Meanwhile, although the CMNs-connected soybean plants did not directly receive Cd supply, they upregulated transcriptional levels of Cd transport-related genes HATPase and RSTK 2.13- and 5.96-fold, respectively, induced activities of POD by 44.8% in the leaves, and increased MDA by 146.2% in the roots. Furthermore, Cd addition inhibited maize growth but mycorrhizal colonization improved plant performance in presence of Cd stress. This finding demonstrates that mycorrhizal networks mediate the transfer of Cd between plants of different species, suggesting a potential to use CMNs as a conduit to transfer toxic heavy metals from main food crops to heavy metal hyperaccumulators via intercropping.

LncRNA TLR8-AS1 promotes metastasis and chemoresistance of ovarian cancer through enhancing TLR8 mRNA stability.

Ovarian cancer is diagnosed as the most deadly gynecological tumor. Ovarian cancer metastasis affects chemoresistance and confers poor patient prognosis. In present work, we intended to elucidate whether long non-coding RNAs (lncRNAs) TLR8-AS1 regulated cell metastasis and chemoresistance of ovarian cancer, and uncover the molecular mechanism of TLR8-AS1 in the modulation of ovarian cancer progression. Firstly, bioinformatics analyses identified TLR8-AS1 as a cancer-associated fibroblasts regulated lncRNA in ovarian cancer. Further experiments revealed that TLR8-AS1 augmented cell metastasis and chemoresistance of ovarian cancer in vitro and in vivo. Moreover, TLR8-AS1 upregulates TLR8 by stabilizing TLR8 mRNA, thus activating NF-κB signaling and promoting ovarian cancer metastasis and chemoresistance. Besides, TCGA data analysis suggested that TLR8-AS1 is elevated in ovarian cancer in comparison to adjacent non-cancerous tissues. High TLR8-AS1 expression levels were measured in metastatic ovarian cancer and correlated with poor patient prognosis. The clinical data supported the mechanism and biological significance of TLR8-AS1 dysregulation in ovarian cancer development. Our work demonstrates that TLR8-AS1 can be applied as a diagnostic and prognostic indicator for ovarian cancer, and maybe an alternative target for the treatment of ovarian cancer.

A dual substrate-accessing mechanism of a major facilitator superfamily protein facilitates lysophospholipid flipping across the cell membrane.

Lysophospholipid transporter (LplT) is a member of the major facilitator superfamily present in many Gram-negative bacteria. LplT catalyzes flipping of lysophospholipids (LPLs) across the bacterial inner membrane, playing an important role in bacterial membrane homeostasis. We previously reported that LplT promotes both uptake of exogenous LPLs and intramembranous LPL flipping across the bilayer. To gain mechanistic insight into this dual LPL-flipping activity, here we implemented a combination of computational approaches and LPL transport analyses to study LPL binding of and translocation by LplT. Our results suggest that LplT translocates LPLs through an elongated cavity exhibiting an extremely asymmetric polarity. We found that two D(E)N motifs form a head group-binding site, in which the carboxylate group of Asp-30 is important for LPL head group recognition. Substitutions of residues in the head group-binding site disrupted both LPL uptake and flipping activities. However, alteration of hydrophobic residues on the interface between the N- and C-terminal domains impaired LPL flipping specifically, resulting in LPLs accumulation in the membrane, but LPL uptake remained active. These results suggest a dual substrate-accessing mechanism, in which LplT recruits LPLs to its substrate-binding site via two routes, either from its extracellular entry or through a membrane-embedded groove between transmembrane helices, and then moves them toward the inner membrane leaflet. This LPL-flipping mechanism is likely conserved in many bacterial species, and our findings illustrate how LplT adjusts the major facilitator superfamily translocation pathway to perform its versatile lipid homeostatic functions.

The phospholipid-repair system LplT/Aas in Gram-negative bacteria protects the bacterial membrane envelope from host phospholipase A2 attack.

Secretory phospholipases A2 (sPLA2s) are potent components of mammalian innate-immunity antibacterial mechanisms. sPLA2 enzymes attack bacteria by hydrolyzing bacterial membrane phospholipids, causing membrane disorganization and cell lysis. However, most Gram-negative bacteria are naturally resistant to sPLA2 Here we report a novel resistance mechanism to mammalian sPLA2 in Escherichia coli, mediated by a phospholipid repair system consisting of the lysophospholipid transporter LplT and the acyltransferase Aas in the cytoplasmic membrane. Mutation of the lplT or aas gene abolished bacterial lysophospholipid acylation activity and drastically increased bacterial susceptibility to the combined actions of inflammatory fluid components and sPLA2, resulting in bulk phospholipid degradation and loss of colony-forming ability. sPLA2-mediated hydrolysis of the three major bacterial phospholipids exhibited distinctive kinetics and deacylation of cardiolipin to its monoacyl-derivative closely paralleled bacterial death. Characterization of the membrane envelope in lplT- or aas-knockout mutant bacteria revealed reduced membrane packing and disruption of lipid asymmetry with more phosphatidylethanolamine present in the outer leaflet of the outer membrane. Moreover, modest accumulation of lysophospholipids in these mutant bacteria destabilized the inner membrane and rendered outer membrane-depleted spheroplasts much more sensitive to sPLA2 These findings indicated that LplT/Aas inactivation perturbs both the outer and inner membranes by bypassing bacterial membrane maintenance mechanisms to trigger specific interfacial activation of sPLA2 We conclude that the LplT/Aas system is important for maintaining the integrity of the membrane envelope in Gram-negative bacteria. Our insights may help inform new therapeutic strategies to enhance host sPLA2 antimicrobial activity.

Substrate Selectivity of Lysophospholipid Transporter LplT Involved in Membrane Phospholipid Remodeling in Escherichia coli.

Lysophospholipid transporter (LplT) was previously found to be primarily involved in 2-acyl lysophosphatidylethanolamine (lyso-PE) recycling in Gram-negative bacteria. This work identifies the potent role of LplT in maintaining membrane stability and integrity in the Escherichia coli envelope. Here we demonstrate the involvement of LplT in the recycling of three major bacterial phospholipids using a combination of an in vitro lysophospholipid binding assay using purified protein and transport assays with E. coli spheroplasts. Our results show that lyso-PE and lysophosphatidylglycerol, but not lysophosphatidylcholine, are taken up by LplT for reacylation by acyltransferase/acyl-acyl carrier protein synthetase on the inner leaflet of the membrane. We also found a novel cardiolipin hydrolysis reaction by phospholipase A2 to form diacylated cardiolipin progressing to the completely deacylated headgroup. These two distinct cardiolipin derivatives were both translocated with comparable efficiency to generate triacylated cardiolipin by acyltransferase/acyl-acyl carrier protein synthetase, demonstrating the first evidence of cardiolipin remodeling in bacteria. These findings support that a fatty acid chain is not required for LplT transport. We found that LplT cannot transport lysophosphatidic acid, and its substrate binding was not inhibited by either orthophosphate or glycerol 3-phosphate, indicating that either a glycerol or ethanolamine headgroup is the chemical determinant for substrate recognition. Diacyl forms of PE, phosphatidylglycerol, or the tetra-acylated form of cardiolipin could not serve as a competitive inhibitor in vitro. Based on an evolutionary structural model, we propose a "sideways sliding" mechanism to explain how a conserved membrane-embedded α-helical interface excludes diacylphospholipids from the LplT binding site to facilitate efficient flipping of lysophospholipid across the cell membrane.

Structural Insight into Substrate Selection and Catalysis of Lipid Phosphate Phosphatase PgpB in the Cell Membrane.

PgpB belongs to the lipid phosphate phosphatase protein family and is one of three bacterial integral membrane phosphatases catalyzing dephosphorylation of phosphatidylglycerol phosphate (PGP) to generate phosphatidylglycerol. Although the structure of its apo form became recently available, the mechanisms of PgpB substrate binding and catalysis are still unclear. We found that PgpB was inhibited by phosphatidylethanolamine (PE) in a competitive mode in vitro Here we report the crystal structure of the lipid-bound form of PgpB. The structure shows that a PE molecule is stabilized in a membrane-embedded tunnel formed by TM3 and the "PSGH" fingerprint peptide near the catalytic site, providing structural insight into PgpB substrate binding mechanism. Noteworthy, in silico docking of varied lipid phosphates exhibited similar substrate binding modes to that of PE, and the residues in the lipid tunnel appear to be important for PgpB catalysis. The catalytic triad in the active site is essential for dephosphorylating substrates lysophosphatidic acid, phosphatidic acid, or sphingosine-1-phosphate but surprisingly not for the native substrate PGP. Remarkably, residue His-207 alone is sufficient to hydrolyze PGP, indicating a specific catalytic mechanism for PgpB in PG biosynthesis. We also identified two novel sensor residues, Lys-93 and Lys-97, on TM3. Our data show that Lys-97 is essential for the recognition of lyso-form substrates. Modification at the Lys-93 position may alter substrate specificity of lipid phosphate phosphatase proteins in prokaryotes versus eukaryotes. These studies reveal new mechanisms of lipid substrate selection and catalysis by PgpB and suggest that the enzyme rests in a PE-stabilized state in the bilayer.

A restriction-free method for gene reconstitution using two single-primer PCRs in parallel to generate compatible cohesive ends.

BACKGROUND: Restriction-free (RF) cloning, a PCR-based method for the creation of custom DNA plasmids, allows for the insertion of any sequence into any plasmid vector at any desired position, independent of restriction sites and/or ligation. Here, we describe a simple and fast method for performing gene reconstitution by modified RF cloning. RESULTS: Double-stranded inserts and acceptors were first amplified by regular PCR. The amplified fragments were then used as the templates in two separate linear amplification reactions containing either forward or reverse primer to generate two single-strand reverse-complement counterparts, which could anneal to each other. The annealed inserts and acceptors with 5' and 3' cohesive ends were sealed by ligation reaction. Using this method, we made 46 constructs containing insertions of up to 20 kb. The average cloning efficiency was higher than 85%, as confirmed by colony PCR and sequencing of the inserts. CONCLUSIONS: Our method provides an alternative cloning method capable of inserting any DNA fragment of up to at least 20 kb into a plasmid, with high efficiency. This new method does not require restriction sites or alterations of the plasmid or the gene of interest, or additional treatments. The simplicity of both primer design and the procedure itself makes the method suitable for high-throughput cloning and structural genomics.

AFEAP cloning: a precise and efficient method for large DNA sequence assembly.

BACKGROUND: Recent development of DNA assembly technologies has spurred myriad advances in synthetic biology, but new tools are always required for complicated scenarios. Here, we have developed an alternative DNA assembly method named AFEAP cloning (Assembly of Fragment Ends After PCR), which allows scarless, modular, and reliable construction of biological pathways and circuits from basic genetic parts. METHODS: The AFEAP method requires two-round of PCRs followed by ligation of the sticky ends of DNA fragments. The first PCR yields linear DNA fragments and is followed by a second asymmetric (one primer) PCR and subsequent annealing that inserts overlapping overhangs at both sides of each DNA fragment. The overlapping overhangs of the neighboring DNA fragments annealed and the nick was sealed by T4 DNA ligase, followed by bacterial transformation to yield the desired plasmids. RESULTS: We characterized the capability and limitations of new developed AFEAP cloning and demonstrated its application to assemble DNA with varying scenarios. Under the optimized conditions, AFEAP cloning allows assembly of an 8 kb plasmid from 1-13 fragments with high accuracy (between 80 and 100%), and 8.0, 11.6, 19.6, 28, and 35.6 kb plasmids from five fragments at 91.67, 91.67, 88.33, 86.33, and 81.67% fidelity, respectively. AFEAP cloning also is capable to construct bacterial artificial chromosome (BAC, 200 kb) with a fidelity of 46.7%. CONCLUSIONS: AFEAP cloning provides a powerful, efficient, seamless, and sequence-independent DNA assembly tool for multiple fragments up to 13 and large DNA up to 200 kb that expands synthetic biologist's toolbox.

Biogenesis, transport and remodeling of lysophospholipids in Gram-negative bacteria.

Lysophospholipids (LPLs) are metabolic intermediates in bacterial phospholipid turnover. Distinct from their diacyl counterparts, these inverted cone-shaped molecules share physical characteristics of detergents, enabling modification of local membrane properties such as curvature. The functions of LPLs as cellular growth factors or potent lipid mediators have been extensively demonstrated in eukaryotic cells but are still undefined in bacteria. In the envelope of Gram-negative bacteria, LPLs are derived from multiple endogenous and exogenous sources. Although several flippases that move non-glycerophospholipids across the bacterial inner membrane were characterized, lysophospholipid transporter LplT appears to be the first example of a bacterial protein capable of facilitating rapid retrograde translocation of lyso forms of glycerophospholipids across the cytoplasmic membrane in Gram-negative bacteria. LplT transports lyso forms of the three bacterial membrane phospholipids with comparable efficiency, but excludes other lysolipid species. Once a LPL is flipped by LplT to the cytoplasmic side of the inner membrane, its diacyl form is effectively regenerated by the action of a peripheral enzyme, acyl-ACP synthetase/LPL acyltransferase (Aas). LplT-Aas also mediates a novel cardiolipin remodeling by converting its two lyso derivatives, diacyl or deacylated cardiolipin, to a triacyl form. This coupled remodeling system provides a unique bacterial membrane phospholipid repair mechanism. Strict selectivity of LplT for lyso lipids allows this system to fulfill efficient lipid repair in an environment containing mostly diacyl phospholipids. A rocker-switch model engaged by a pair of symmetric ion-locks may facilitate alternating substrate access to drive LPL flipping into bacterial cells. This article is part of a Special Issue entitled: Bacterial Lipids edited by Russell E. Bishop.

The Melibiose Transporter of Escherichia coli: CRITICAL CONTRIBUTION OF LYS-377 TO THE STRUCTURAL ORGANIZATION OF THE INTERACTING SUBSTRATE BINDING SITES.

We examine the role of Lys-377, the only charged residue in helix XI, on the functional mechanism of the Na(+)-sugar melibiose symporter from Escherichia coli. Intrinsic fluorescence, FRET, and Fourier transform infrared difference spectroscopy reveal that replacement of Lys-377 with either Cys, Val, Arg, or Asp disables both Na(+) and melibiose binding. On the other hand, molecular dynamics simulations extending up to 200-330 ns reveal that Lys-377 (helix XI) interacts with the anionic side chains of two of the three putative ligands for cation binding (Asp-55 and Asp-59 in helix II). When Asp-59 is protonated during the simulations, Lys-377 preferentially interacts with Asp-55. Interestingly, when a Na(+) ion is positioned in the Asp-55-Asp-59 environment, Asp-124 in helix IV (a residue essential for melibiose binding) reorients and approximates the Asp-55-Asp-59 pair, and all three acidic side chains act as Na(+) ligands. Under these conditions, the side chain of Lys-377 interacts with the carboxylic moiety of these three Asp residues. These data highlight the crucial role of the Lys-377 residue in the spatial organization of the Na(+) binding site. Finally, the analysis of the second-site revertants of K377C reveals that mutation of Ile-22 (in helix I) preserves Na(+) binding, whereas that of melibiose is largely abolished according to spectroscopic measurements. This amino acid is located in the border of the sugar-binding site and might participate in sugar binding through apolar interactions.

The substitution of Arg149 with Cys fixes the melibiose transporter in an inward-open conformation.

The melibiose transporter from Escherichia coli (MelB) can use the electrochemical energy of either H(+), Na(+) or Li(+) to transport the disaccharide melibiose to the cell interior. By using spectroscopic and biochemical methods, we have analyzed the role of Arg149 by mutagenesis. According to Fourier transform infrared difference and fluorescence spectroscopy studies, R149C, R149Q and R149K all bind substrates in proteoliposomes, where the protein is disposed inside-out. Analysis of right-side-out (RSO) and inside-out (ISO) membrane vesicles showed that the functionally active R149Q and R149K mutants could bind externally added fluorescent sugar analog in both types of vesicles. In contrast, the non-transporting R149C mutant does bind the fluorescent sugar analog as well as melibiose and Na(+) in ISO, but not in RSO vesicles. Therefore, the mutation of Arg149 into cysteine restrains the orientation of transporter to an inward-open conformation, with the inherent consequences of a) reducing the frequency of access of outer substrates to the binding sites, and b) impairing active transport. It is concluded that Arg149, most likely located in the inner (cytoplasmic) half of transmembrane helix 5, is critically involved in the reorientation mechanism of the substrate-binding site accessibility in MelB.

An NFAT1-C3a-C3aR Positive Feedback Loop in Tumor-Associated Macrophages Promotes a Glioma Stem Cell Malignant Phenotype.

Extensive infiltration by tumor-associated macrophages (TAM) in combination with myeloid-derived suppressor cells constitute the immunosuppressive microenvironment and promote the malignant phenotype of gliomas. The aggressive mesenchymal (MES)-subtype glioma stem cells (GSC) are prominent in the immunosuppressive microenvironment of gliomas. However, the underlying immune-suppressive mechanisms are still unknown. The current study showed that the antitumor immune microenvironment was activated in glioma in Nfat1-/- mice, suggesting induction of the immune-suppressive microenvironment by nuclear factor of activated T cells-1 (NFAT1). In TAMs, NFAT1 could upregulate the transcriptional activity of complement 3 (C3) and increase the secretion of C3a, which could then bind to C3aR and promote M2-like macrophage polarization by activating TIM-3. Simultaneously, C3a/C3aR activated the Ca2+-NFAT1 pathway, forming a positive feedback loop for the M2-like polarization of TAMs, which further promoted the MES transition of GSCs. Finally, disruption of this feedback loop using a C3aR inhibitor significantly inhibited glioma growth both in vitro and in vivo. The current study demonstrated that a NFAT1-C3a-C3aR positive feedback loop induces M2-like TAMs and further promotes the malignant phenotype of GSCs, which might be the potential therapeutic target for glioma.

Protein Dynamics Mediated by Cardiolipin in Bacteria.

Bacterial proteins targeting the appropriate subcellular sites are the base for their proper function. Several studies have shown that the anionic phospholipid cardiolipin (CL), a conical lipid preferring negative membrane curvature, modulates the lipid bilayers' structure, which impacts the activity of their resident proteins. Due to the favor of negative membrane curvature, CL is not randomly distributed in the bacterial plasma membrane. In contrast, it gathers in particular parts of the cell membrane to form microdomains, in which many functional membrane proteins are accumulated and carry out diverse physiological processes of bacteria, such as cell division, metabolism, infection, and antibiotic residence. In addition, CL has a unique structure that carries two negative charges, which makes it play a pivotal role in protein assembly, interaction, and location. These characteristics of CL make it closely related to many crucial physiological functions of bacteria. Here, we have reviewed the mechanism of protein dynamics mediated by CL initiated on the bacterial membrane. Furthermore, we studied the effect of CL on bacterial infection and antibiotic residence. Finally, the CL-targeting therapeutic agents for antibacterial therapy are also examined.

Genomic Insights into the Pathogenicity and Drug-Resistance of a Bacillus cereus Isolated from Human Teeth.

Background: Bacillus cereus is a common bacterium found in the environment. Some strains can cause food poisoning, and very few can cause clinically severe infections, leading to death. Here, we characterized the genome sequence of B. cereus LIN78 isolated from teeth with deep caries and compared it with those of 25 other related species. Methods: Third-generation sequencing technology, bacteriological analyses, biochemistry, and mass spectrometry were applied to characterize the drug-resistance genes and virulence factors of B. cereus LIN78. Results: The complete genome sequence of B. cereus Lin78 consists of 5647 genes distributed on a circular chromosome, a 393 kbp plasmid, and 928 pseudogenes (37.4% of whole-genome DNA). The LIN78 genome contains 14 sets of 16s, 23s, and 5s ribosomal RNA operons; 106 tRNA genes, one tmRNA, 12 genomic islands, six prophases, 64 repeats; 37 antibiotic-resistant genes; and 1119 putative virulence genes, including enterotoxins and cytolysins. The B. cereus LIN78 genome carries multiple copies of non-ribosomal polypeptide synthetase (NRPS) and post-translationally modified peptides (RiPPs). Phylogenetic analysis of the 26 B. cereus strains showed that B. cereus LIN78 is evolutionarily closely related to B. thuringiensis ATCC 10792 and B. cereus ATCC 14579. Conclusion: The newly isolated B. cereus carries many virulence genes, including enterotoxins and hemolysins, similar to B. anthracis, and multiple antibiotic resistance genes. These findings suggest that the strain has a potential risk of causing disease. Our studies are vital for further exploration of the evolution of B. cereus, its pathogenic mechanisms, and the control and treatment of bacterial infections.

CCL2 mediated IKZF1 expression promotes M2 polarization of glioma-associated macrophages through CD84-SHP2 pathway.

The proneural-mesenchymal (PN-MES) transformation of glioma stem cells (GSCs) can significantly increase proliferation, invasion, chemotherapy tolerance, and recurrence. M2-like polarization of tumor-associated macrophages (TAMs) has a strong immunosuppressive effect, promoting tumor malignancy and angiogenesis. There is limited understanding on the interactions between GSCs and TAMs as well as their associated molecular mechanisms. In the present study, bioinformatics analysis, GSC and TAM co-culture, determination of TAM polarization phenotypes, and other in vitro experiments confirmed that CCL2 secreted by MES-GSCs promotes TAM-M2 polarization via the IKZF1-CD84-SHP2 pathway and PN-MES transformation of GSCs via the IKZF1-LRG1 pathway in TAMs. IKZF1 inhibitors could significantly reduce tumor volumes in animal glioma models and improve survival, as well as suppress TAM-M2 polarization and the GSC malignant phenotype. The results of this study indicate the important interaction between TAMs and GSCs in the glioma microenvironment as well as its role in tumor progression. The findings also suggest a novel target for follow-up clinical transformation research on the regulation of TAM function and GSCs malignant phenotype.

Genomic Insights into the First Emergence of blaNDM-5-Carrying Carbapenem-Resistant Salmonella enterica Serovar London Strain in China.

Carbapenem-resistant Salmonella enterica (S. enterica) pose a significant threat to public health, causing gastroenteritis and invasive infections. We report the first emergence of a carbapenem-resistant S. enterica serovar London strain, A132, carrying the blaNDM-5 gene in China. Whole-genome sequencing and bioinformatics analysis assigned A132 to be ST155, a multidrug-resistant clone frequently reported in China. The strain A132 exhibited resistance to multiple antibiotics, with 20 acquired antibiotic resistance genes (ARGs) identified, predominantly located on the IncFIB plasmid (pA132-1-NDM). Notably, the blaNDM-5 gene was located within an IS26 flanked-class 1 integron-ISCR1 complex, comprising two genetic cassettes. One cassette is the class 1 integron, which may facilitate the transmission of the entire complex, while the other is the blaNDM-5-containing ISCR1-IS26-flanked cassette, carrying multiple other ARGs. Genbank database search based on the blaNDM-5-carrying cassette identified a similar genetic context found in transmissible IncFIA plasmids from Escherichia coli (p91) and Enterobacter hormaechei (p388) with a shared host range, suggesting the potential for cross-species transmission of blaNDM-5. To our knowledge, this is the first reported case of Salmonella serovar London ST155 harboring blaNDM-5 gene. Phylogenetic analysis indicated a close relationship between A132 and eight S. London ST155 strains isolated from the same province. However, A132 differed by carrying the blaNDM-5 gene and four unique ARGs. Given the high transmissibility of the F-type plasmid harboring blaNDM-5 and 18 other ARGs, it is imperative to implement vigilant surveillance and adopt appropriate infection control measures to mitigate the threat to public health.

Adhesive polydopamine-based photothermal hybrid hydrogel for on-demand lidocaine delivery, effective anti-bacteria, and prolonged local long-lasting analgesia.

Considering the astonishing prevalence of localized pain affecting billions of patients worldwide, the development of advanced analgesic formulations or delivery systems to achieve clinical applicability is of great significance. In this study, an integrated PDA-based LiH@PDA@Ag@PAA@Gelatin system was designed for sustained delivery of lidocaine hydrochloride (LiH). By optimizing the preparation process and formulation of the hydrogel, the hydrogel exhibited superior mechanical properties, reversibility, adhesion strength, and self-healing attributes. Moreover, PDA@Ag nanoparticles were evenly dispersed within the hydrogel, and the optimized PDA@Ag@PAA@Gelatin showed a higher photothermal conversion efficiency than that of pure PDA. Importantly, LiH@PDA@Ag@PAA@Gelatin could effectively capture and eradicate bacteria through the synergistic interaction between near-infrared (NIR), PDA, Ag and LiH. In vitro and in vivo tests demonstrated that LiH@PDA@Ag@PAA@Gelatin exhibited higher drug delivery efficiency compared to commercial lidocaine patches. By evaluating the mechanical pain withdrawal threshold of the spared nerve injury (SNI) model in rats, it was proven that LiH@PDA@Ag@PAA@Gelatin enhanced and prolonged the analgesic effect of LiH. Furthermore, LiH@PDA@Ag@PAA@Gelatin induced by NIR possessed excellent on-demand photothermal analgesic ability. Therefore, this study develops a convenient method for preparing localized analgesic hydrogel patches, providing an important step towards advancing PDA-based on-demand pain relief applications.