Salmonella spvC gene suppresses macrophage/neutrophil antibacterial defense mediated by gasdermin D.

OBJECTIVE: Salmonella enterica serovar Typhimurium (S. Typhimurium) is a representative model organism for investigating host-pathogen interactions. It was reported that S. Typhimurium spvC gene alleviated intestinal inflammation to aggravate systemic infection, while the precise mechanisms remain unclear. In this study, the influence of spvC on the antibacterial defense of macrophage/neutrophil mediated by gasdermin D (GSDMD) was investigated. METHODS: Mouse macrophage-like cell lines J774A.1 and RAW264.7, neutrophil-like cells derived from HL-60 cells (human promyletic leukemia cell lines) were infected with S. Typhimurium wild type, spvC deletion and complemented strains. Cell death was evaluated by LDH release and Annexin V-FITC/PI staining. Macrophage pyroptosis and neutrophil NETosis were detected by western blotting, live cell imaging and ELISA. Flow cytometry was used to assess the impact of spvC on macrophage-neutrophil cooperation in macrophage (dTHP-1)-neutrophil (dHL-60) co-culture model pretreated with GSDMD inhibitor disulfiram. Wild-type and Gsdmd-/- C57BL/6J mice were utilized for in vivo assay. The degree of phagocytes infiltration and inflammation were analyzed by immunofluorescence and transmission electron microscopy. RESULTS: Here we find that spvC inhibits pyroptosis in macrophages via Caspase-1/Caspase-11 dependent canonical and non-canonical pathways, and restrains neutrophil extracellular traps extrusion in GSDMD-dependent manner. Moreover, spvC could ameliorate macrophages/neutrophils infiltration and cooperation in the inflammatory response mediated by GSDMD to combat Salmonella infection. CONCLUSIONS: Our findings highlight the antibacterial activity of GSDMD in phagocytes and reveal a novel pathogenic mechanism employed by spvC to counteract this host defense, which may shed new light on designing effective therapeutics to control S. Typhimurium infection.

Effects of sericin and egg white on the inflammation of damaged skin in mice.

Sericin and egg white (EW) have shown the ability to promote wound healing. However, there have been insufficient studies regarding the effects of sericin and EW mixtures on wound healing. This study aimed to investigate the effects of a hybrid sericin and EW solution on wound repair and inflammation-related indicators in mouse skin. In this work, sericin with a low molecular weight was first mixed with homogeneous EW to prepare a hybrid wound dressing. Histology evaluation, the expression of C-reactive protein (CRP) and inflammatory cytokines in mice were tested to determine the effects of this dressing on skin injuries in mice. The results showed that sericin and the hybrid solution of sericin and EW effectively promoted wound healing in mouse skin. The wound recovery rates of mice 12 days after treatment with a medium dose of sericin (0.2 g ml-1) and the same dosage of sericin with added EW were 1.32 and 1.65 times that of mice treated with phosphate buffer saline as a control, respectively. In addition, the mixture solution was more effective in wound healing than sericin alone. Sericin with EW significantly reduced the expression of CRP and inflammatory cytokines in mice during wound healing. A sericin and EW hybrid solution can effectively shorten the time needed for wound healing and reduce inflammation-related indicators in mice, making it a promising candidate for wound dressing.

Evaluation of Thermal Fatigue Life and Crack Morphology in Brake Discs of Low-Alloy Steel for High-Speed Trains.

Effective braking in high-speed trains is one of the major bottlenecks in expediting the technology and possibilities to improve speed. Although substantial progress has been made to increase operating speed, perhaps, thermal fatigue cracking in brake discs is a primary constraint so far. Thermal fatigue cracking is the major cause of brake disc failure in high-speed trains, especially trains with a speed of 350 km/h or above. In this study, new material composition is proposed for brake discs of high-speed trains. A comprehensive investigation is presented based on fatigue crack initiation and propagation, along with wear and micro-hardness characterization. Thermal fatigue tests at various thermal cycles between 20 ℃ and 700 ℃ were performed and the experimental results are compared with fatigue properties of a commercial brake disc material. An experimental trial revealed that thermal cracks normally initiate and propagate along the oxidized grain boundaries; nevertheless, crack propagation is restricted by the fine precipitates and lath structure of martensitic. Moreover, crack length at the initiation and propagation stage is predicted through crack growth rate and favorable grain size in the crack vicinity. Thermal fatigue life can be improved by dictating the microstructure and precipitate morphology of cast steel by tailoring the alloying composition.

Prevalence and Antimicrobial Resistance of Ureaplasma urealyticum and Mycoplasma hominis in Patients with Genital Tract Infection in Jiangsu, China.

BACKGROUND: The aim of this study was to investigate the infection and antimicrobial resistance of Ureaplasma urealyticum (U. urealyticum) and Mycoplasma hominis (M. hominis) in patients with genital tract diseases in Jiangsu, China. METHODS: A total of 3,321 patients suspected with genital tract infectious diseases were enrolled in this study from September 2017 to September 2020. The Mycoplasma detection and antimicrobial susceptibility were tested using the commercially available Mycoplasma kit. RESULTS: Among the 3,321 specimens tested, 1,503 (45.3%) were positive for Mycoplasmas, and the proportion of mono-infection of U. urealyticum is highest (79.5%). The overall infection rate has been increasing in the past 3 years. The positive rate in females (68.7%) was higher than in males (25.0%), and the main infection age group was 20 - 39 (81.2%). Besides, U. urealyticum and M. hominis displayed relative lower resistance rates to gatifloxacin, josamycin, minocycline, and doxycycline (6.0%, 6.5%, 3.1%, and 3.2%, respectively). However, the antimicrobial resistance rates to azithromycin, clindamycin, roxithromycin, sparfloxacin, and ofloxacin were relatively high (45.4%, 42.1%, 34.9, 36.0, and 65.5%, respectively). Antimicrobial resistance of U. urealyticum and M. hominis to these 14 drugs have been changing in the past 3 years. CONCLUSIONS: In total, these preliminary data showed the prevalence and antimicrobial resistance status of U. urealyticum and M. hominis in patients suspected with genital tract infectious diseases, which has use for reference on both prevention and treatment of diseases caused by them.

Salmonella spvC Gene Inhibits Autophagy of Host Cells and Suppresses NLRP3 as Well as NLRC4.

Salmonella spvC gene, encoding a phosphothreonine lyase on host mitogen-activated protein kinases, facilitates systemic infection of Salmonella while the precise mechanisms remain elusive. Autophagy and pyroptosis dependent on the activation of inflammasomes, as parts of innate immune response, contribute to host defense against Salmonella infection. Recently, we reported that spvC could inhibit pyroptosis. To explore the effect of spvC on autophagy and the relationship between its function in pyroptosis and autophagy, infection models of macrophages J774A.1 and epithelial HeLa cells co-cultured with Salmonella Typhimurium wild type, spvC deletion, site-directed mutant which lacks phosphothreonine lyase activity, or complemented strain were established. The levels of LC3 turnover and Beclin 1 of J774A.1 cells were determined by western blot. Confocal laser scanning microscopy was used to visualize the autophagic flux after being transfected with mRFP-GFP-LC3 plasmid in HeLa cells. Results showed that SpvC inhibited autophagosome formation through its phosphothreonine lyase activity. Additionally, analysis of nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) and NLR with CARD domain-containing 4 (NLRC4) in J774A.1 cells indicated that spvC decreased the protein levels of NLRP3 and NLRC4, which were significantly changed by autophagy inhibitor Bafilomycin A1. Together, our observations reveal a novel mechanism of spvC in Salmonella pathogenesis and host inflammatory response via inhibiting autophagy and NLRP3 as well as NLRC4. These pathways and their subversion by diverse pathogen virulence determinants are expected to throw light on the design of anti-infective agents.

Salmonella spvC Gene Inhibits Pyroptosis and Intestinal Inflammation to Aggravate Systemic Infection in Mice.

Salmonella enterica serovar Typhimurium (S). Typhimurium is a primary foodborne pathogen infecting both humans and animals. Salmonella plasmid virulence C (spvC) gene is closely related to S. Typhimurium dissemination in mice, while the mechanisms remain to be fully elucidated. Pyroptosis, a gasdermin-mediated inflammatory cell death, plays a role in host defense against bacterial infection, whereas the effect of spvC on pyroptosis and its function in inflammatory injury induced by S. Typhimurium are rather limited. In our study, C57BL/6 mice and J774A.1 cells infected with S. Typhimurium wild-type strain SL1344, spvC deletion mutant, spvC K136A site-directed mutant, and complemented strain were used to investigate potential pathogenesis of spvC. We verity that SpvC attenuates intestinal inflammation, suppresses pyroptosis through phosphothreonine lyase activity, and reduces pyroptosis in the ceca. Moreover, the reduction of inflammation via spvC results in systemic infection. These findings demonstrate that spvC inhibits pyroptosis and intestinal inflammation to promote bacterial dissemination, which provide new strategies for controlling systemic infection caused by Salmonella and novel insights for the treatment of other corresponding diseases.

Salmonella spv locus affects type I interferon response and the chemotaxis of neutrophils via suppressing autophagy.

Salmonella is a facultative intracellular pathogen that can cause significant morbidity and mortality in humans and animals. Salmonella plasmid virulence (spv) gene sequence is a highly conserved 6.8 kb region which exists in the plasmid of most pathogenic Salmonella. Autophagy is a degradation process of unnecessary and dysfunctional cytoplasm components to maintain cellular homeostasis, which could affect host inflammatory responses, such as type I interferon response. Type I interferon response can promote the antibacterial activity of macrophage as well as the secretion of cytokines and neutrophil chemokines. We previously reported that spv locus could suppress autophagy and the aggregation of neutrophils in zebrafish larvae. To explore the influence of spv locus on Salmonella escaping from the innate immune responses and the underlying mechanism, the models of Salmonella enterica serovar Typhimurium infected macrophages in vitro and zebrafish larvae in vivo were used in this study. The interactions among spv locus, autophagy, type I interferon response and the chemotaxis of neutrophils were investigated. Western blot was used to detect the expression levels of autophagy related proteins and RT-qPCR was used to measure the mRNA levels of type I interferon response and the neutrophil chemokines. The chemotaxis of neutrophils were observed by Laser Scanning confocal microscopy. Autophagy agonist Torin 1 was also involved to interfere the autophagy influx. Results showed that spv locus could restrain type I interferon response and the chemotaxis of neutrophils via suppressing autophagy, which provided substantial foundation to study the mechanism of Salmonella escaping the innate immunity.