GRB7-mediated enhancement of cell malignant characteristics induced by Helicobacter pylori infection.

Growth factor receptor bound protein 7 (GRB7) is reportedly upregulated in human gastric cancer (GC), which is closely associated with tumor progression and prognosis. However, the mechanism underlying its dysregulation in GC remains poorly understood. In this study, we found that GRB7 overexpression was associated with Helicobacter pylori (H. pylori) infection. GC cells (AGS and MGC-803) infection assays revealed that this upregulation was mediated by the transcription factor STAT3, and activation of STAT3 by H. pylori promoted GRB7 expression in infected GC cells. Moreover, CagA, the key virulence factor of H. pylori, was found involved in STAT3-mediated GRB7 overexpression. The overexpressed GRB7 further promoted GC cell proliferation, migration, and invasion by activating ERK signaling. Mice infection was further used to investigate the action of GRB7. In H. pylori infection, GRB7 expression in mice gastric mucosa was elevated, and higher STAT3 and ERK activation were also detected. These results revealed GRB7-mediated pathogenesis in H. pylori infection, in which H. pylori activates STAT3, leading to increased GRB7 expression, then promotes activation of the ERK signal, and finally enhances malignant properties of infected cells. Our findings elucidate the role of GRB7 in H. pylori-induced gastric disorders, offering new prospects for the treatment and prevention of H. pylori-associated gastric carcinogenesis by targeting GRB7.

Integrin-Linked Kinase in the Development of Gastric Tumors Induced by Helicobacter pylori: Regulation and Prevention Potential.

BACKGROUND: Integrin-linked kinase (ILK) is crucial in solid tumors by regulating the Hippo-Yes-associated protein 1 (YAP) pathway. This study aimed to uncover how Helicobacter pylori influences ILK levels and its role in regulating YAP during H. pylori-induced gastric cancer. MATERIALS AND METHODS: GES-1 cells with stable Ilk knockdown and overexpression and a mouse carcinogenesis model for H. pylori infection were constructed. And ILK, the phosphorylated mammalian STE20-like protein kinase 1 (MST1), large tumor suppressor 1 (LATS1; S909, T1079), and YAP (S109, S127) were detected in cells, and mice by western blotting, as well as fluorescence intensity of YAP were assayed by immunofluorescence. YAP downstream genes Igfbp4 and Ctgf, the pathological changes and tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), interleukin-1beta (IL-1ß), and nitric oxide (NO) levels in mice gastric tissues were detected by real-time PCR, H&E, and ELISA assays. RESULTS: In this study, stable Ilk knockdown cells exhibited significantly higher phosphorylated levels of MST1, LATS1, and YAP, as well as increased YAP in the nuclei of GES-1 cells. Conversely, cells with Ilk overexpression showed opposite results. H. pylori infection led to decreased ILK levels in gastric epithelial cells but increased ILK levels in gastric cancer cell lines (MGC803, SGC7901) and gastric cancer tissues in mice. Treatment with the ILK inhibitor OST-T315 elevated the phosphorylated MST, LATS1, and YAP levels, and inhibited the mRNA levels of Igfbp4 and Ctgf at 44, 48 week-aged mice. OST-T315 also reduced the release of TNF-α, IL-6, IL-1ß, and NO, as well as the progression of gastric cancer caused by H. pylori and N-Nitroso-N-methylurea (NMU) treatment. CONCLUSION: Upon initiation of gastric tumorigenesis signals, H. pylori increases ILK levels and suppresses Hippo signaling, thereby promoting YAP activation and gastric cancer progression. ILK can serve as a potential prevention target to impede H. pylori-induced gastric cancer.

Helicobacter pylori East Asian type CagA hijacks more SHIP2 by its EPIYA-D motif to potentiate the oncogenicity.

CagA is a significant oncogenic factor injected into host cells by Helicobacter pylori, which is divided into two subtypes: East Asian type (CagAE), characterized by the EPIYA-D motif, and western type (CagAW), harboring the EPIYA-C motif. CagAE has been reported to have higher carcinogenicity than CagAW, although the underlying reason is not fully understood. SHIP2 is an intracellular phosphatase that can be recruited by CagA to perturb the homeostasis of intracellular signaling pathways. In this study, we found that SHIP2 contributes to the higher oncogenicity of CagAE. Co-Immunoprecipitation and Pull-down assays showed that CagAE bind more SHIP2 than CagAW. Immunofluorescence staining showed that a higher amount of SHIP2 recruited by CagAE to the plasma membrane catalyzes the conversion of PI(3,4,5)P3 into PI(3,4)P2. This alteration causes higher activation of Akt signaling, which results in enhanced IL-8 secretion, migration, and invasion of the infected cells. SPR analysis showed that this stronger interaction between CagAE and SHIP2 stems from the higher affinity between the EPIYA-D motif of CagAE and the SH2 domain of SHIP2. Structural analysis revealed the crucial role of the Phe residue at the Y + 5 position in EPIYA-D. After mutating Phe of CagAE into Asp (the corresponding residue in the EPIYA-C motif) or Ala, the activation of downstream Akt signaling was reduced and the malignant transformation of infected cells was alleviated. These findings revealed that CagAE hijacks SHIP2 through its EPIYA-D motif to enhance its carcinogenicity, which provides a better understanding of the higher oncogenic risk of H. pylori CagAE.

Antagonizing roles of SHP1 in the pathogenesis of Helicobacter pylori infection.

BACKGROUND: SHP1 has been documented as a tumor suppressor and it was thought to play an antagonistic role in the pathogenesis of Helicobacter pylori infection. In this study, the exact mechanism of this antagonistic action was studied. MATERIALS AND METHODS: AGS, MGC803, and GES-1 cells were infected with H. pylori, intracellular distribution changes of SHP1 were first detected by immunofluorescence. SHP1 overexpression and knockdown were then constructed in these cells to investigate its antagonistic roles in H. pylori infection. Migration and invasion of infected cells were detected by transwell assay, secretion of IL-8 was examined via ELISA, the cells with hummingbird-like alteration were determined by microexamination, and activation of JAK2/STAT3, PI3K/Akt, and ERK pathways were detected by immunoblotting. Mice infection model was established and gastric pathological changes were evaluated. Finally, the SHP1 activator sorafenib was used to analyze the attenuating effect of SHP1 activation on H. pylori pathogenesis in vitro and in vivo. RESULTS: The sub-localization of SHP1 changed after H. pylori infection, specifically that the majority of the cytoplasmic SHP1 was transferred to the cell membrane. SHP1 inhibited H. pylori-induced activation of JAK2/STAT3 pathway, PI3K/Akt pathway, nuclear translocation of NF-κB, and then reduced EMT, migration, invasion, and IL-8 secretion. In addition, SHP1 inhibited the formation of CagA-SHP2 complex by dephosphorylating phosphorylated CagA, reduced ERK phosphorylation and the formation of CagA-dependent hummingbird-like cells. In the mice infection model, gastric pathological changes were observed and increased IL-8 secretion, indicators of cell proliferation and EMT progression were also detected. By activating SHP1 with sorafenib, a significant curative effect against H. pylori infection was obtained in vitro and in vivo. CONCLUSIONS: SHP1 plays an antagonistic role in H. pylori pathogenesis by inhibiting JAK2/STAT3 and PI3K/Akt pathways, NF-κB nuclear translocation, and CagA phosphorylation, thereby reducing cell EMT, migration, invasion, IL-8 secretion, and hummingbird-like changes.

Chronic exposure to polystyrene nanoplastics induces intestinal mechanical and immune barrier dysfunction in mice.

Micro(nano)plastics are prevalent in the environment, and prolonged exposure to them represents a threat to human health. The goal of this study is to assess the health risk of long-term exposure to nanoplastics (NPs) at environmental concentrations on the intestinal mechanical and immune barrier in mice. In this study, mice were provided drinking water containing polystyrene NPs (PS-NPs; 0.1, 1, and 10 mg·L-1) for 32 consecutive weeks. The levels of endocytosis proteins caveolin and clathrin and of tight junctional proteins claudin-1, occludin, and ZO-1, and morphological changes, proportion of lymphocytes B in MLNs and lymphocytes T in IELs and LPLs were determined by immunohistochemistry, hematoxylin-eosin, and flow cytometry assays in the intestinal tissues of mice at 28 weeks. The activities or concentrations of ROS, SOD, MDA, and GSH-Px and inflammatory factors (IL-1ß, IL-6, and TNF-α) in the intestinal tissues of mice were measured by ELISA at 12, 16, 20, 24, and 32 weeks. Compared with the control group, oral ingested PS-NPs entered the intestinal tissues of mice and upregulated expression levels of the clathrin and caveolin. The intestinal tissue structure of mice in the PS-NPs (1 and 10 mg·L-1) exposure groups showed significant abnormalities, such as villus erosion, decreased of crypts numbers and large infiltration of inflammatory cells. Exposure to 0.1 mg·L-1 PS-NPs decreased occludin protein levels, but not claudin-1 and ZO-1 levels. The levels of these three tight junction proteins decreased significantly in the 1 and 10 mg·L-1 PS-NPs exposed groups. Exposure to PS-NPs led to a significant time- and dose-dependent increase in ROS and MDA levels, and concurrently decreased GSH-Px and SOD contents. Exposure to PS-NPs increased the proportion of B cells in MLNs, and decreased the proportion of CD8+ T cells in IELs and LPLs. The levels of pro-inflammatory cytokines IL-6, TNF-α and IL-1ß were markedly elevated after PS-NPs exposure. Long-term PS-NPs exposure impaired intestinal mechanical and immune barrier, and indicate a potentially significant threat to human health.

Chloroquine inhibited Helicobacter pylori-related gastric carcinogenesis by YAP-ß-catenin-autophagy axis.

YAP participates in autophagy associated with many diseases. In this study, we demonstrate that YAP promotes autophagy by interacting with beclin 1, upregulating beclin 1 and LC3B-II protein expression, and promoting autophagosome formation after H. pylori infection in a vacuolating cytotoxin A-dependent manner. The protein levels of ß-catenin in the cytoplasm and nuclei of GES-1 cells and the mRNA levels of Axin2, Myc, Lgr5, and Ccnd1 were increased in H. pylori-infected cells or YAP-overexpressed cells, but were decreased in YAP-silenced cells. The ß-catenin inhibitor XAV939 significantly downregulated autophagy, whereas the activator LiCl showed opposite effects. An H. pylori-infected mouse model of gastric carcinoma was successfully established. The mouse model showed that H. pylori infection, when combined with NMU, promoted the tumorigenesis of gastric tissues; increased IL-1ß, IL-6, and TNF-α levels; promoted NO release; and increased the expression of beclin 1, LC3B-II more than NMU alone. Chloroquine inhibited these phenomena, but did not completely attenuate the effects of H. pylori. These results demonstrate that chloroquine can be used as a drug for the treatment of H. pylori-related gastric cancer, but the treatment should simultaneously remove H. pylori.

Helicobacter pylori infection induces autophagy via ILK regulation of NOXs-ROS-Nrf2/HO-1-ROS loop.

Reactive oxygen species (ROS) can regulate the occurrence of autophagy, and effective control of the balance between ROS and autophagy may be an important strategy for Helicobacter pylori induced gastric-related diseases. In this study, infection with H. pylori led to a lower level of ILK phosphorylation and increased ROS generation. Knockdown of ILK enhanced total ROS generation, and upregulated NADPH oxidase (NOX) subunit p22-phox levels. Inhibition of NOXs affected total ROS generation. The inhibition of NOX and ROS generation reduced Nrf2 and HO-1 levels, and knockdown of ILK significantly enhanced Nrf2 levels in H. pylori-infected GES-1 cells. Activation of Nrf2 by DMF decreased ROS levels. Therefore, NOX-dependent ROS production regulated by ILK was essential for activation of Nrf2/HO-1 signaling pathways in H. pylori-infected GES-1 cells. Beclin1, ATG5 and LC3B-II levels were higher both in H. pylori-infected and ILK-knockdown GES-1 cells. In NAC-pretreated GES-1 cells infected with H. pylori, the LC3B-II level was decreased compared to that in cells after H. pylori infection alone. Stable low expression of ILK with further knockdown of Beclin1 or ATG5 significantly reduced LC3B-II levels in GES-1 cells, while with the addition of the autophagy inhibitor chloroquine (CQ), LC3B-II and p62 protein levels were both remarkably upregulated. H. pylori accelerated the accumulation of ROS and further led to the induction of ROS-mediated autophagy by inhibiting ILK levels. Together, these results indicate that H. pylori infection manipulates the NOX-ROS-Nrf2/HO-1-ROS loop to control intracellular oxygen stress and further induced ROS-mediated autophagy by inhibiting ILK levels.

The phosphotransferase system gene ptsH affects persister formation in Klebsiella pneumoniae by regulating cyclic adenosine monophosphate levels.

Klebsiella pneumoniae is one of the most common opportunistic pathogens causing hospital- and community-acquired infections. Antibiotic resistance in K. pneumoniae has emerged as a major clinical and public health threat. Persisters are specific antibiotic-tolerant bacterial cells. Studies on the mechanism underlying their formation mechanism and growth status are scarce. Therefore, it is urgent to explore the key genes and signalling pathways involved in the formation and recovery process of K. pneumoniae persisters to enhance the understanding and develop relevant treatment strategies. In this study, we treated K. pneumoniae with a lethal concentration of levofloxacin. It resulted in a distinct plateau of surviving levofloxacin-tolerant persisters. Subsequently, we obtained bacterial samples at five different time points during the formation and recovery of K. pneumoniae persisters to perform transcriptome analysis. ptsH gene was observed to be upregulated during the formation of persisters, and down-regulated during the recovery of the persisters. Further, we used CRISPR-Cas9 to construct ΔptsH, the ptsH-knockout K. pneumoniae strain, and to investigate the effect of ptsH on the persister formation. We observed that ptsH can promote the formation of persisters, reduce accumulation of reactive oxygen species, and enhance antioxidant capacity by reducing cyclic adenosine monophosphate (cAMP) levels. To the best of our knowledge, this is the first study to report that ptsH plays a vital role in forming K. pneumoniae persisters. This study provided important insights to further explore the mechanism underlying the formation of K. pneumoniae persisters and provided a potential target for treating infection with K. pneumoniae persisters.

Analysis of cross neutralizing activity of antibodies from sera of severe fever with thrombocytopenia syndrome patients to deal with different genotype strains.

Background: Severe fever with thrombocytopenia syndrome bunyavirus (SFTSV) is a tick-borne virus that causes severe communicable fever with thrombocytopenia syndrome (SFTS) with an average case fatality rate of 10%. In the study, we aimed to identify the cross-neutralizing antibody (nAb) against different genotype strains from sera of SFTSV infected patients. Methods: Firstly the genotype of SFTSV was identified by constructing a phylogenetic tree based on the M segments epidemic in the Jiaodong area of Shandong province, then different sera of subjects cross reactive with recombinant Gn (rGn-Fc) or recombinant Gc (rGc-Fc) of 0921 strain were examined. The levels of polyclonal nAbs from sera of 25 convalescents were measured by a pseudovirus-based neutralizing experiment. Results: We found local endemic strains were mainly C2 and C3 isolates of SFTSV. 14 of 15 sera from donors reacted with 0921 rGn-Fc, and 9 of 15 sera from donors reacted with 0921 rGc-Fc. Cross nAbs were produced by 10 of 25 sera from donors during the period of 2019-2021. Among these, five nAbs (A2, A4, A5, L9, and L10) neutralized the pseudoviruses of HB29, Gangwon, HN13, HN20, SPL030A, and SD4 strains. Conclusion: Our data suggested that epidemic strains showed relatively stable heredity. Some blood sources from patients produced cross nAbs that could neutralize all of the strains examined. These findings highlight the important role played by humoral immunity in combatting SFTSV.

Oceanospirillum sediminis sp. nov., Isolated From Coastal Sediment in the Yellow Sea.

A novel Gram-negative, motile, aerobic, spiral-shaped bacterium designated D5T, was isolated from a coastal sediment collected in the Yellow Sea. Optimal growth occurred at 30 °C, pH 7.0-8.0 and in the presence of 1-3% (w/v) NaCl. Strain D5T contained ubiquinone 8 (Q-8) as the predominant respiratory quinone. The major fatty acids (> 10%) were C16:0, C16:1 ω7c/C16:1 ω6c and C18:1w7c/C18:1w6c. The main polar lipids were phosphatidylglycerol and phosphatidylethanolamine. The draft genome is 5.6 Mb in length, and DNA G + C content is 47.2 mol%. 16S rRNA gene sequences showed that strain D5T is most closely related to Oceanospirillum beijerinckii NBRC 15445T (97.8%, sequence similarity). However, the digital DNA-DNA hybridization (dDDH) value and average nucleotide identity (ANI) between strain D5T and O. beijerinckii is only 27.8% and 77.1%. Phylogenetic trees based on 16S rRNA gene sequences and whole genomes all indicated that strain D5T formed a separate branch in the genus Oceanospirillum. Combined results of the polyphasic analyses suggested that strain D5T represents a novel species in the genus Oceanospirillum, for which the name Oceanospirillum sediminis sp. nov. is proposed. The type strain is D5T (= MCCC 1K06061T = KCTC 62987T).