Production of α-ketoisovalerate with whey powder by systemic metabolic engineering of Klebsiella oxytoca.

BACKGROUND: Whey, which has high biochemical oxygen demand and chemical oxygen demand, is mass-produced as a major by-product of the dairying industry. Microbial fermentation using whey as the carbon source may convert this potential pollutant into value-added products. This study investigated the potential of using whey powder to produce α-ketoisovalerate, an important platform chemical. RESULTS: Klebsiella oxytoca VKO-9, an efficient L-valine producing strain belonging to Risk Group 1 organism, was selected for the production of α-ketoisovalerate. The leucine dehydrogenase and branched-chain α-keto acid dehydrogenase, which catalyzed the reductive amination and oxidative decarboxylation of α-ketoisovalerate, respectively, were inactivated to enhance the accumulation of α-ketoisovalerate. The production of α-ketoisovalerate was also improved through overexpressing α-acetolactate synthase responsible for pyruvate polymerization and mutant acetohydroxyacid isomeroreductase related to α-acetolactate reduction. The obtained strain K. oxytoca KIV-7 produced 37.3 g/L of α-ketoisovalerate from lactose, the major utilizable carbohydrate in whey. In addition, K. oxytoca KIV-7 also produced α-ketoisovalerate from whey powder with a concentration of 40.7 g/L and a yield of 0.418 g/g. CONCLUSION: The process introduced in this study enabled efficient α-ketoisovalerate production from low-cost substrate whey powder. Since the key genes for α-ketoisovalerate generation were integrated in genome of K. oxytoca KIV-7 and constitutively expressed, this strain is promising in stable α-ketoisovalerate fermentation and can be used as a chassis strain for α-ketoisovalerate derivatives production.

Innovative nebulization delivery of lipid nanoparticle-encapsulated siRNA: a therapeutic advance for Staphylococcus aureus-induced pneumonia.

BACKGROUND: Integrin α5ß1 plays a crucial role in the invasion of nonphagocytic cells by Staphylococcus aureus (S. aureus), thereby facilitating infection development. Lipid nanoparticles (LNPs) serve as an effective vehicle for delivering small interfering ribonucleic acids (siRNA) that represent a method to knockdown integrin α5ß1 in the lungs through nebulization, thereby potentially mitigating the severity of S. aureus pneumonia. The aim of this study was to harness LNP-mediated targeting to precisely knockdown integrin α5ß1, thus effectively addressing S. aureus-induced pneumonia. METHODS: C57 mice (8 week-old females) infected with S. aureus via an intratracheal nebulizing device were utilized for the experiments. The LNPs were synthesized via microfluidic mixing and characterized by their size, polydispersity index, and encapsulation efficiency. Continuous intratracheal nebulization was employed for consistent siRNA administration, with the pulmonary function metrics affirming biosafety. The therapeutic efficacy of LNP-encapsulated siRNAs against pneumonia was assessed through western blotting, bacterial count measurement, quantitative polymerase chain reaction, and histological analyses. RESULTS: LNPs, which have an onion-like structure, retained integrity post-nebulization, ensuring prolonged siRNA stability and in vivo safety. Intratracheal nebulization delivery markedly alleviated the severity of S. aureus-induced pneumonia, as indicated by reduced bacterial load and bolstered immune response, thereby localizing the infection to the lungs and averting systemic dissemination. CONCLUSIONS: Intratracheal nebulization of LNP-encapsulated siRNAs targeting integrin α5ß1 significantly diminished the S. aureus-mediated cellular invasion and disease progression in the lungs, presenting a viable therapeutic approach for respiratory infections.

Organoids: new frontiers in tumor immune microenvironment research.

The tumor microenvironment (TME) contains cells that regulate medication response and cancer growth in a major way. Tumor immunology research has been rejuvenated and cancer treatment has been changed by immunotherapy, a rapidly developing therapeutic approach. The growth patterns of tumor cells in vivo and the heterogeneity, complexity, and individuality of tumors produced from patients are not reflected in traditional two-dimensional tumor cell profiles. On the other hand, an in vitro three-dimensional (3D) model called the organoid model is gaining popularity. It can replicate the physiological and pathological properties of the original tissues in vivo. Tumor cells are the source of immune organoids. The TME characteristics can be preserved while preserving the variety of tumors by cultivating epithelial tumor cells with various stromal and immunological components. In addition to having genetic and physical similarities to human diseases and the ability to partially reconstruct the complex structure of tumors, these models are now widely used in research fields including cancer, developmental biology, regenerative mechanisms, drug development, disease modeling, and organ transplantation. This study reviews the function of organoids in immunotherapy and the tumor immune milieu. We also discuss current developments and suggest translational uses of tumor organoids in immuno-oncology research, immunotherapy modeling, and precision medicine.

An Ultrasensitive Biosensor for Probing Subcellular Distribution and Mitochondrial Transport of l-2-Hydroxyglutarate.

l-2-Hydroxyglutarate (l-2-HG) is a functionally compartmentalized metabolite involved in various physiological processes. However, its subcellular distribution and mitochondrial transport remain unclear owing to technical limitations. In the present study, an ultrasensitive l-2-HG biosensor, sfLHGFRH, composed of circularly permuted yellow fluorescent protein and l-2-HG-specific transcriptional regulator, is developed. The ability of sfLHGFRH to be used for analyzing l-2-HG metabolism is first determined in human embryonic kidney cells (HEK293FT) and macrophages. Then, the subcellular distribution of l-2-HG in HEK293FT cells and the lower abundance of mitochondrial l-2-HG are identified by the sfLHGFRH-supported spatiotemporal l-2-HG monitoring. Finally, the role of the l-glutamate transporter SLC1A1 in mitochondrial l-2-HG uptake is elucidated using sfLHGFRH. Based on the design of sfLHGFRH, another highly sensitive biosensor with a low limit of detection, sfLHGFRL, is developed for the point-of-care diagnosis of l-2-HG-related diseases. The accumulation of l-2-HG in the urine of patients with kidney cancer is determined using the sfLHGFRL biosensor.

The Role of Hydrogel Biomaterials in the Intervention of Wound Healing and Skin Regeneration via Exosomes: A Systematic Review and Meta-Analysis of Preclinical Animal Studies.

Significance: The combination of hydrogel biomaterials with exosomes to facilitate wound healing and skin regeneration is a promising approach. Recent Advances: Recent preclinical animal studies have focused on investigating the efficacy of hydrogel-based delivery systems for exosomes in promoting wound healing and skin regeneration. Critical Issues: Despite encouraging results, critical issues remain unresolved, such as optimizing hydrogel properties to enhance the efficacy of combined therapy with exosomes for wound and bridging the translational gap between preclinical and clinical applications. Future Directions: Future research endeavors should concentrate on refining hydrogel design to enhance exosome delivery efficacy, conducting rigorous clinical trials to assess the safety and efficacy of exosome-loaded hydrogels in human wound healing and skin regeneration, and exploring innovative strategies to maximize therapeutic outcomes.

Identification and Characterization of Bacterial Alliinase: Resource and Substrate Stereospecificity.

Limited alliinase resources cause difficulties in the biosynthesis of thiosulfinates (e.g., allicin), restricting their applications in the agricultural and food industries. To effectively biosynthesize thiosulfinates, this study aimed to excavate bacterial alliinase resources and elucidate their catalytic properties. Two bacterial cystathionine ß-lyases (MetCs) possessing high alliinase activity (>60 U mg -1) toward L-(-)-alliin were identified from Allium sativum rhizosphere isolates. Metagenomic exploration revealed that cystathionine ß-lyase from Bacillus cereus (BcPatB) possessed high activity toward both L-(±)-alliin and L-(+)-alliin (208.6 and 225.1 U mg -1), respectively. Although these enzymes all preferred l-cysteine S-conjugate sulfoxides as substrates, BcPatB had a closer phylogenetic relationship with Allium alliinases and shared several similar features with A. sativum alliinase. Interestingly, the Trp30Ile31Ala32Asp33 Met34 motif in a cuspate loop of BcPatB, especially sites 31 and 32 at the top of the motif, was modeled to locate near the sulfoxide of L-(+)-alliin and is important for substrate stereospecificity. Moreover, the stereoselectivity and activity of mutants I31V and A32G were higher toward L-(+)-alliin than those of mutant I31L/D33E toward L-(-)-alliin. Using bacterial alliinases and chemically synthesized substrates, we obtained thiosulfinates with high antimicrobial and antinematode activities that could provide insights into the protection of crops and food.

IFN-α affects Th17/Treg cell balance through c-Maf and associated with the progression of EBV- SLE.

OBJECTIVES: Systemic lupus erythematosus (SLE) is a multi-organ autoimmune disease, of which the pathogens is remains obscure. Viral infection, particularly Epstein Barr viru (EBV) infection, has been considered a common pathogenic factor. This study suggests that c-Maf may be an important target in T cell differentiation during SLE progression, providing a potentially new perspective on the role of viral infection in the pathogenesis of autoimmune diseases. METHODS: Cytokines of EBV-infected SLE patients were measured by ELISA and assessed in conjunction with their clinical data. IFN-α, c-Maf, and the differentiation of Th17/Treg cells in SLE patients and MRL/LPR mice were analyzed using FCM, WB, RT-PCR, etc. Following the infection of cells and mice with EBV or viral mimic poly (dA:dT), the changes of the aforementioned indicators were investigated. The relationship among IFN-α, STAT3, c-Maf and Th17 cells was determined by si-RNA technique. RESULTS: Many SLE patients are found to be complicated by viral infections; Further, studies have demonstrated that viral infection, especially EBV, is involved in SLE development. This study showed that viral infections might promote IFN-α secretion, inhibit c-Maf expression by activating STAT3, increase Th17 cell differentiation, and lead to the immune imbalance of Th17/Treg cells, thus playing a role in the onset and progression of SLE. CONCLUSION: This study demonstrates that EBV infections may contribute to SLE development by activating STAT3 through IFN-α, inhibiting c-Maf, and causing Th17/Treg immune imbalance. Our work provided a new insight into the pathogenesis and treatment of SLE.

TRIM38 Induced in Respiratory Syncytial Virus-infected Cells Downregulates Type I Interferon Expression by Competing with TRIM25 to Bind RIG-I.

Innate immune response is the first line of defense for the host against virus invasion. One important response is the synthesis and secretion of type I interferon (IFN-I) in the virus-infected host cells. Here, we found that respiratory syncytial virus (RSV) infection induced high expression of TRIM25, which belongs to the tripartite motif-containing (TRIM) family of proteins. TRIM25 bound and activated retinoic acid-inducible gene I (RIG-I) by K63-linked ubiquitination. Accordingly, RIG-I mediated the production of IFN-I mainly through the nuclear factor kappa-B (NF-κB) pathway in respiratory epithelial cells. Interestingly, IFN-I, in turn, promoted a high expression of TRIM38 which downregulated the expression of IFN-I by reducing the protein level of RIG-I by K48-linked ubiquitination. More importantly, the binding site of TRIM25 to RIG-I was found in the narrow 25th-43rd amino acid (aa) region of RIG-I N-terminus. In contrast, the binding sites of TRIM38 to RIG-I were found in a much wider amino acid region, which included the binding site of TRIM25 on RIG-I. As a result, TRIM38 inhibits the production of IFN-I by competing with TRIM25 for RIG-I binding. Thus, TRIM38 negatively regulates RIG-I activation to, in turn, downregulate IFN-I expression, thus interfering with host immune response. A negative feedback loop effectively "puts the brakes" on the reaction once host immune response is overactivated and homeostasis is unbalanced. We also discovered that TRIM25 bound RIG-I by a new K63-linked ubiquitination located at K-45 of the first caspase recruitment domain (CARD). Collectively, these results confirm an antagonism between TRIM38 and TRIM25 in regulating IFN-I production by affecting RIG-I activity following RNA virus infection.

Advances and Applications of Cancer Organoids in Drug Screening and Personalized Medicine.

In recent years, the rapid emergence of 3D organoid technology has garnered significant attention from researchers. These miniature models accurately replicate the structure and function of human tissues and organs, offering more physiologically relevant platforms for cancer research. These intricate 3D structures not only serve as promising models for studying human cancer, but also significantly contribute to the advancement of various potential applications in the field of cancer research. To date, organoids have been efficiently constructed from both normal and malignant tissues originating from patients. Using such bioengineering platforms, simulations of infections and cancer processes, mutations and carcinogenesis can be achieved, and organoid technology is also expected to facilitate drug testing and personalized therapies. In conclusion, regenerative medicine has the potential to enhance organoid technology and current transplantation treatments by utilizing genetically identical healthy organoids as substitutes for irreversibly deteriorating diseased organs. This review explored the evolution of cancer organoids and emphasized the significant role these models play in fundamental research and the advancement of personalized medicine in oncology.

Efficient production of 1,2,4-butanetriol from corn cob hydrolysate by metabolically engineered Escherichia coli.

Corn cob is a major waste mass-produced in corn agriculture. Corn cob hydrolysate containing xylose, arabinose, and glucose is the hydrolysis product of corn cob. Herein, a recombinant Escherichia coli strain BT-10 was constructed to transform corn cob hydrolysate into 1,2,4-butanetriol, a platform substance with diversified applications. To eliminate catabolite repression and enhance NADPH supply for alcohol dehydrogenase YqhD catalyzed 1,2,4-butanetriol generation, ptsG encoding glucose transporter EIICBGlc and pgi encoding phosphoglucose isomerase were deleted. With four heterologous enzymes including xylose dehydrogenase, xylonolactonase, xylonate dehydratase, α-ketoacid decarboxylase and endogenous YqhD, E. coli BT-10 can produce 36.63 g/L 1,2,4-butanetriol with a productivity of 1.14 g/[L·h] using xylose as substrate. When corn cob hydrolysate was used as the substrate, 43.4 g/L 1,2,4-butanetriol was generated with a productivity of 1.09 g/[L·h] and a yield of 0.9 mol/mol. With its desirable characteristics, E. coli BT-10 is a promising strain for commercial 1,2,4-butanetriol production.

Rab5c promotes RSV and ADV replication by autophagy in respiratory epithelial cells.

Respiratory system diseases caused by respiratory viruses are common and exert tremendous pressure on global healthcare system. In our previous studies, we found that Long non-coding RNA NRAV (Lnc NRAV) and its target molecule Rab5c plays a significant role in respiratory virus infection. However, the mechanism by which Rab5c affects virus replication remains unclear. Rab5c, a protein mainly localized on the cell membranes and in early endosomes and phagosomes, participates in endocytosis mediated by clathrin and regulates the fusion of early endosome, maturation of early phagosomes, and autophagy. Therefore, we inferred that Rab5c impacts virus replication, which might be related to endocytosis or autophagy. We selected RSV (respiratory syncytial virus) as a representative enveloped virus and ADV (Adenovirus) as a representative non-enveloped virus to explore the possible mechanism of RSV and ADV replication promoted by Rab5c in A549 cells and in Rab5c-overexpressing mice. Here, we confirmed that the activated Rab5c promotes RSV and ADV replication and the inactivated Rab5c inhibits their replication. However, Rab5c promoting RSV and ADV replication is not mediated by endocytosis rather by autophagy in respiratory epithelial cells. Our study showed that Rab5c upregulates LC3-Ⅱ (microtubule-associated protein 1 light chain 3 beta) protein expression levels by interacting with Beclin1, a key autophagy molecule, which can induce autophagy and promote replication of ADV and RSV. This study enriches the understanding of the interaction between respiratory viruses and Rab5c, providing new insights for virus prevention and treatment.

Promoting microbial fermentation in lignocellulosic hydrolysates by removal of inhibitors using MTES and PEI-modified chitosan-chitin nanofiber hybrid aerogel.

To further enhance the removal efficiency for furanic and phenolic compounds in lignocellulosic hydrolysates, a new detoxification strategy was proposed, which retained fermentable sugars and promoted the growth and metabolism of subsequent bacteria. The best adsorbent (P/M-CCA) was prepared by hybrid chitosan-chitin nanofiber, graft modification with polyethylenimine, and silanization with methyl triethoxylsilane in order. Taken corn cob hydrolysate as object, the removal rates of HMF and furfural were 85.1 % and 99.0 %, respectively. The removal rates of six out of nine phenolic inhibitors were 100 %, and the other three were more than 65 %. Even better, the retention rates of glucose and xylose were both 100 %. In contrast to no growth in undetoxified hydrolysates, Bacillus coagulans grew normally in detoxified hydrolysates, and lactic acid reached 19.1 g/L after 12 h fermentation. P/M-CCA achieves both removal of multiple inhibitors and retain sugars, which would promote the valorization of highly toxic lignocellulosic hydrolysates.

Efficient L-valine production using systematically metabolic engineered Klebsiella oxytoca.

L-Valine, a branched-chain amino acid with diversified applications, is biosynthesized with α-acetolactate as the key precursor. In this study, the metabolic flux in Klebsiella oxytoca PDL-K5, a Risk Group 1 organism producing 2,3-butanediol as the major fermentation product, was rearranged to L-valine production by introducing exogenous L-valine biosynthesis pathway and blocking endogenous 2,3-butanediol generation at the metabolic branch point α-acetolactate. After further enhancing L-valine efflux, strengthening pyruvate polymerization and selecting of key enzymes for L-valine synthesis, a plasmid-free K. oxytoca strain VKO-9 was obtained. Fed-batch fermentation with K. oxytoca VKO-9 in a 7.5 L fermenter generated 122 g/L L-valine with a yield of 0.587 g/g in 56 h. In addition, repeated fed-batch fermentation was conducted to prevent precipitation of L-valine due to oversaturation. The average concentration, yield, and productivity of produced L-valine in three cycles of repeated fed-batch fermentation were 81.3 g/L, 0.599 g/g, and 3.39 g/L/h, respectively.

A biosensor for D-2-hydroxyglutarate in frozen sections and intraoperative assessment of IDH mutation status.

The oncometabolite D-2-hydroxyglutarate (D-2-HG) has emerged as a valuable biomarker in tumors with isocitrate dehydrogenase (IDH) mutations. Efficient detection methods are required and rapid intraoperative determination of D-2-HG remains a huge challenge. Herein, D-2-HG dehydrogenase from Achromobacter xylosoxidans (AX-D2HGDH) was found to have high substrate specificity. AX-D2HGDH dehydrogenizes D-2-HG and reduces flavin adenine dinucleotide (FAD) bound to the enzyme. Interestingly, the dye resazurin can be taken as another substrate to restore FAD. AX-D2HGDH thus catalyzes a bisubstrate and biproduct reaction: the dehydrogenation of D-2-HG to 2-ketoglutarate and simultaneous reduction of non-fluorescent resazurin to highly fluorescent resorufin. According to steady-state analysis, a ping-pong bi-bi mechanism has been concluded. The Km values for resazurin and D-2-HG were determined as 0.56 µM and 10.93 µM, respectively, suggesting high affinity to both substrates. On the basis, taking AX-D2HGDH and resazurin as recognition and fluorescence transducing element, a D-2-HG biosensor (HGAXR) has been constructed. HGAXR exhibits high sensitivity, accuracy and specificity for D-2-HG in different biological samples. With the aid of HGAXR and the matched low-cost palm-size detecting device, D-2-HG levels in frozen sections of resected brain tumor tissues can be measured in a direct, simple and accurate manner with a fast detection (1-3 min). As the technique of frozen section is familiar to surgeons and pathologists, HGAXR and the portable device can be easily integrated into the current workflow, having potential to provide rapid intraoperative pathology for IDH mutation status and guide decision-making during surgery.

ß-Galactosidase: a traditional enzyme given multiple roles through protein engineering.

ß-Galactosidases are crucial carbohydrate-active enzymes that naturally catalyze the hydrolysis of galactoside bonds in oligo- and disaccharides. These enzymes are commonly used to degrade lactose and produce low-lactose and lactose-free dairy products that are beneficial for lactose-intolerant people. ß-galactosidases exhibit transgalactosylation activity, and they have been employed in the synthesis of galactose-containing compounds such as galactooligosaccharides. However, most ß-galactosidases have intrinsic limitations, such as low transglycosylation efficiency, significant product inhibition effects, weak thermal stability, and a narrow substrate spectrum, which greatly hinder their applications. Enzyme engineering offers a solution for optimizing their catalytic performance. The study of the enzyme's structure paves the way toward explaining catalytic mechanisms and increasing the efficiency of enzyme engineering. In this review, the structure features of ß-galactosidases from different glycosyl hydrolase families and the catalytic mechanisms are summarized in detail to offer guidance for protein engineering. The properties and applications of ß-galactosidases are discussed. Additionally, the latest progress in ß-galactosidase engineering and the strategies employed are highlighted. Based on the combined analysis of structure information and catalytic mechanisms, the ultimate goal of this review is to furnish a thorough direction for ß-galactosidases engineering and promote their application in the food and dairy industries.

The nonstructural protein 1 of respiratory syncytial virus hijacks host mitophagy as a novel mitophagy receptor to evade the type I IFN response in HEp-2 cells.

Mitochondria are good targets for viruses to manipulate their hosts. However, it remains obscure whether respiratory syncytial virus (RSV) target mitochondria to suppress the type I interferon (IFN) responses. Here, we show that nonstructural protein 1 (NS1) protein of RSV interacts with Tu translation elongation factor mitochondrial (TUFM), which can lead to its localization in mitochondria and finally induce TUFM-dependent mitophagy and inhibition of IFNß. Mechanically, NS1-mediated TUFM-dependent mitophagy does not depend on the PINK1-PARKIN pathway and classic mitophagy receptors. Importantly, NS1 may act as a new receptor protein to bridge mitochondria and autophagosomes by interacting with TUFM and LC3B. The LIR motif of NS1 protein is essential for its interaction with LC3B and is of great importance for its mitophagy induction and IFNß suppression. Finally, NS1-induced TUFM-dependent mitophagy was essential for its attenuated IFNß response using autophagy-deficient cells and mice. Our study provides a novel mitophagy receptor molecular and a new antiviral option by suppressing antiviral innate immune via targeting TUFM-dependent mitophagy. IMPORTANCE It is a worthy concern for us to understand virus-host interactions which affect progression and prognosis of disease. We demonstrated that the non-structural protein 1 of respiratory syncytial virus (RSV NS1) may act as a novel mitophagy receptor to induce mitophagy by binding LC3B and mitochondrial protein TUFM, and finally dampen interferon (IFN) responses induced by RIG1 and RSV infection. TUFM is beneficial for RSV replication in vivo and vitro. It is new and interesting that RSV NS1 may function as a mitophagy receptor to interact with LC3B. The LIR motif of NS1 protein is essential for its interaction with LC3B. We further confirm that RSV NS1 inhibited IFNß response and promoted RSV replication in autophagy-dependent mechanisms in vivo and vitro. Our study contributes to understanding virus-host interaction, enriching our insights into RSV pathogenic mechanism and exploiting new antiviral treatments targeting TUFM.

Poly-3-hydroxybutyrate production from acetate by recombinant Pseudomonas stutzeri with blocked L-leucine catabolism and enhanced growth in acetate.

Acetate is a low-cost feedstock for the production of different bio-chemicals. Electrochemical reduction of CO2 into acetate and subsequent acetate fermentation is a promising method for transforming CO2 into value-added chemicals. However, the significant inhibitory effect of acetate on microbial growth remains a barrier for acetate-based biorefinery. In this study, the deletion of genes involved in L-leucine degradation was found to be beneficial for the growth of Pseudomonas stutzeri A1501 in acetate. P. stutzeri (Δpst_3217), in which the hydroxymethylglutaryl-CoA lyase catalyzing ß-hydroxy-ß-methylglutaryl-CoA into acetyl-CoA and acetoacetate was deleted, grew faster than other mutants and exhibited increased tolerance to acetate. Then, the genes phbCAB from Ralstonia eutropha H16 for poly-3-hydroxybutyrate (PHB) biosynthesis were overexpressed in P. stutzeri (∆pst_3217) and the recombinant strain P. stutzeri (∆pst_3217-phbCAB) can accumulate 0.11 g L-1 PHB from commercial acetate. Importantly, P. stutzeri (∆pst_3217-phbCAB) can also use CO2-derived acetate to produce PHB and the accumulated PHB accounted for 5.42% (w/w) of dried cell weight of P. stutzeri (∆pst_3217-phbCAB).

Streptococcus pyogenes: Pathogenesis and the Current Status of Vaccines.

Streptococcus pyogenes (group A Streptococcus; GAS), a Gram-positive coccal bacterium, poses a significant global disease burden, especially in low- and middle-income countries. Its manifestations can range from pharyngitis and skin infection to severe and aggressive diseases, such as necrotizing fasciitis and streptococcal toxic shock syndrome. At present, although GAS is still sensitive to penicillin, there are cases of treatment failure for GAS pharyngitis, and antibiotic therapy does not universally prevent subsequent disease. In addition to strengthening global molecular epidemiological surveillance and monitoring of antibiotic resistance, developing a safe and effective licensed vaccine against GAS would be the most effective way to broadly address GAS-related diseases. Over the past decades, the development of GAS vaccines has been stalled, mainly because of the wide genetic heterogeneity of GAS and the diverse autoimmune responses to GAS. With outbreaks of scarlet fever in various countries in recent years, accelerating the development of a safe and effective vaccine remains a high priority. When developing a GAS vaccine, many factors need to be considered, including the selection of antigen epitopes, avoidance of self-response, and vaccine coverage. Given the challenges in GAS vaccine development, this review describes the important virulence factors that induce disease by GAS infection and how this has influenced the progression of vaccine development efforts, focusing on several candidate vaccines that are further along in development.

Irisin Ameliorates PM2.5-Induced Acute Lung Injury by Regulation of Autophagy Through AMPK/mTOR Pathway.

Background: PM2.5 exposure is one of the major inducements of various respiratory diseases and related mortality. Meanwhile, irisin, a metabolism and thermogenesis-related hormone, is found to be protective against acute lung injury induced by LPS, which indicates its therapeutic function in lung injury. However, the function and underlying mechanism of irisin in PM2.5-induced acute lung injury (ALI) are still unclear. This study is aimed to discover the potential mechanisms of irisin in PM2.5-induced acute lung injury. Methods: Atg5 deficient mice and cells were established to clarify the relationship between irisin and autophagy in PM2.5-induced ALI. We also used Ad-mCherry-GFP-LC3B as a monitor of autophagy flux to claim the effects of irisin on autophagy. Western blotting and qPCR were used to reveal the molecular mechanism. Results: As a result, PM2.5 exposure induced lung injury whereas mitigated by irisin. Moreover, PM2.5 hampered autophagy flux, characterized by accumulation of p62, and autophagosomes, as well as blocked autolysosomes. Irisin improved the disturbed autophagy flux, which was abrogated by deficiency of Atg5. Additionally, we demonstrated that irisin activated AMPK and inhibited mTOR, which indicated the enhanced autophagy. Moreover, blockage of AMPK by compound C terminated irisin's induction of autophagy in cultured MH-S cells. Conclusion: Our findings reveal that irisin performs protective effects against PM2.5-induced ALI by activating autophagy through AMPK/mTOR signaling pathway.

Use of machine learning-based integration to develop a monocyte differentiation-related signature for improving prognosis in patients with sepsis.

BACKGROUND: Although significant advances have been made in intensive care medicine and antibacterial treatment, sepsis is still a common disease with high mortality. The condition of sepsis patients changes rapidly, and each hour of delay in the administration of appropriate antibiotic treatment can lead to a 4-7% increase in fatality. Therefore, early diagnosis and intervention may help improve the prognosis of patients with sepsis. METHODS: We obtained single-cell sequencing data from 12 patients. This included 14,622 cells from four patients with bacterial infectious sepsis and eight patients with sepsis admitted to the ICU for other various reasons. Monocyte differentiation trajectories were analyzed using the "monocle" software, and differentiation-related genes were identified. Based on the expression of differentiation-related genes, 99 machine-learning combinations of prognostic signatures were obtained, and risk scores were calculated for all patients. The "scissor" software was used to associate high-risk and low-risk patients with individual cells. The "cellchat" software was used to demonstrate the regulatory relationships between high-risk and low-risk cells in a cellular communication network. The diagnostic value and prognostic predictive value of Enah/Vasp-like (EVL) were determined. Clinical validation of the results was performed with 40 samples. The "CBNplot" software based on Bayesian network inference was used to construct EVL regulatory networks. RESULTS: We systematically analyzed three cell states during monocyte differentiation. The differential analysis identified 166 monocyte differentiation-related genes. Among the 99 machine-learning combinations of prognostic signatures constructed, the Lasso + CoxBoost signature with 17 genes showed the best prognostic prediction performance. The highest percentage of high-risk cells was found in state one. Cell communication analysis demonstrated regulatory networks between high-risk and low-risk cell subpopulations and other immune cells. We then determined the diagnostic and prognostic value of EVL stabilization in multiple external datasets. Experiments with clinical samples demonstrated the accuracy of this analysis. Finally, Bayesian network inference revealed potential network mechanisms of EVL regulation. CONCLUSIONS: Monocyte differentiation-related prognostic signatures based on the Lasso + CoxBoost combination were able to accurately predict the prognostic status of patients with sepsis. In addition, low EVL expression was associated with poor prognosis in sepsis.