Merged yellow dots indicate autophagosomes, and autolysosome numbers were calculated as number of red dots minus number of yellow dots. A Representative images of the xenophagy state after different treatments for 12 h. B Quantitative results of autophagosomes. C Quantitative results of autolysosomes. Effects of PA on H. A Representative images of xenophagy after treatment with gentamycin and H. The results indicate that PA improves the xenophagic flux and promotes digestion.
The expression of miRc-3p, miRc-5p, and miRb-5p was remarkably upregulated by H. The results revealed that H. B Quantitative gene expression results of miRc-3p, miRc-5p, and miRb-5p.
Assays of lysosome activity Figure 8A indicated that H. The transfection of miRb-5p to the cell nucleus remarkably increased 6, 12, and 24 h after H. Effects of PA on lysosome function. A Effects of PA on H. The increased resistance of H. Indeed, the resistance rate of the pathogen to metronidazole, clarithromycin, and levofloxacin has been reported to be as high as The invasion of H. Therefore, sufficient first-time treatment is crucial. The rates of secondary resistance of H. Our results of the comparative study indicated that some clinical strains have stronger invasive ability than standard strains, and the intracellular invasion may be a survival strategy of H.
Another research revealed that the invasive ratio of NCTC is 2. To date, whether the intracellular survival of H. And our lab has begun a clinical experiment to explore the influence of increasing invasion of H. In short, these findings strongly suggest that eradication of intracellular H. Intracellular survival contributes to the drug tolerance of H.
The drug-resistant H. Besides, unfavorable environments with low concentrations of antibiotics may promote bacterial invasion. Pretreatment with PA decreased the intracellular number of H. These findings demonstrate the promising potential clinical applications of PA in intracellular bacteria therapy.
Immunofluorescence and gentamycin protection assays were adopted to analyze the intracellular number of H. Immunofluorescence images enable the detailed observation of intracellular bacteria. However, the time-consuming gentamycin protection assay provides more accurate data because dead bacteria can also be stained.
TEM revealed that H. Similar results were obtained in the adenovirus transfection assay, which showed that PA increases the abundance of autolysosomes and promotes the digestion of H.
Previous studies revealed that the VacA secreted by H. The bacteria then hijack the xenophagy process, inhibit autophagosome formation, and reduce cathepsin D in lysosomes, all of which contribute to the intracellular survival of H. MiRNA families play a key role in H. KEGG analysis of cellular processes and genetic information processing was performed on the basis of the results of miRNA chip assay, and findings indicated that PA affects ubiquitin, the endoplasmic reticulum, and autophagy.
Further verification assay confirmed the effects of PA on miRc-3p, miRc-5p, and miRb-5p, and the related target genes. TFEB is the master regulator of autophagy and lysosomal biogenesis and transfers to the nucleus to modulate the autophagic flux Yang et al. However, the role of TFEB in lysosome modulation could be partly blocked by the upregulation of nuclear miRb-5p induced by H.
Collectively, the elimination of lysosome activity by H. On the one hand, H. On the other hand, the upregulation of nuclear miRb-5p blocks the activity of TFEB and inhibits lysosome function.
Although no effect of PA on TFEB translocation was shown, the downregulation of miRb-5p may contribute to the improvement of lysosome function in the studied system. PA, from P. The compound also shows high cellular uptake and transport efficiency and has been demonstrated to exert therapeutic effects against H.
The high cell-membrane permeability of PA renders it as an effective agent against intracellular H. Our data revealed the bactericidal effects of PA on intracellular H. This study provides an extensive assessment of the multiple therapeutic effects of PA, which may present synergistic effects in combination with other antibiotics against H.
Strains of H. The THP-1 cells were stimulated with H. Cathepsin X expression on THP-1 cells and concentration of cytokines in the supernatant of THP-1 cells were measured with a flow cytometer.
TSS H. We demonstrated a H. Differences in lipid A of H. Marshall and Warren isolated Helicobacter pylori from gastric biopsy samples in Since then, H. The reason for these discrepancies is to be found in the bacterial and host factors that influence the development of the disease [ 2 ]. Recent study in Slovenia estimated the prevalence of H.
Why all patients with H. One of the important host factors that affect cure rates is the immune response to the infection. Activation of the above-mentioned cells leads to H. In the case of impaired host immunity, a defective immune response to H. Chronic exposure to H.
CTSX is mainly found in the cells of the immune system of monocyte lineage, especially macrophages and dendritic cells. Higher levels of CTSX were also found in immune cells of prostate and gastric carcinomas and in macrophages of gastric mucosa infected with H.
It was discovered that patients with H. CTSX was also upregulated in the gastric mucosa of patients with gastric cancer in comparison with those without gastric cancer [ 13 , 14 ]. In our experiments, THP-1 cells, a human monocytic cell line, were used because THP-1 cells are frequently used as a model system for monocytes.
They have been shown to respond with a similar transcriptional pattern as peripheral blood mononuclear cells derived macrophages after stimulation with lipopolysaccharide LPS from Escherichia coli [ 15 , 16 ].
Results from German study suggest that H. We were interested in the differences in the expression of CTSX and cytokine immune response if we stimulate immune cells with H. We included 14 strains of H. All patients needed reevaluation after unsuccessful eradication therapy. Seven patients had H. We used the strains that were isolated from gastric biopsies taken at initial gastroscopy and named them therapy sensitive group—TSS group.
All of these patients eradicated bacteria after second try with the same therapy and the same regime as in the first unsuccessful attempt. Seven patients failed to eradicate the bacteria although they followed advice of a gastroenterologist, how to take the therapy.
The strains were again sensitive to all antibiotics that were used in the first attempt to eradicate H. We used strains of H. These patients failed to eradicate the bacteria till their therapy was switched from amoxicillin plus clarithromycin to amoxicillin plus metronidazole for 14 days.
The switch to the eventually successful therapy was done after second unsuccessful treatment in 5 patients and after third unsuccessful treatment in 2 patients. In these 2 patients H. All patients were well informed about the study, they agreed to have additional gastroscopies, and they documented antibiotic use by specifying day and time of every drug intake. The study was approved by the National Medical Ethics Committee of the Ministry of Health of the Republic of Slovenia, and written consent was obtained from patients before specimen collection.
Antibiotic resistance and minimal inhibitory concentration MIC were determined by E -test. MIC for metronidazole was tested with agar dilution method. Antibiotic susceptibility testing was done on Mueller Hinton agar with old sheep blood.
Antibiotic resistance was tested on all isolates. Typical bacterial colonies were collected, suspended in phosphate buffered saline PBS , pH 7. Suspensions of bacterial antigens were filtered through 0. Cells were incubated in the presence of H.
The plate was then placed on ice and the cells were harvested with ice-cold PBS. In the negative control sample, the antigen was not added to the cells. These are important factors that influence the natural course of H. In our study, all the tested H.
Our results, like those of the previous study, showed that all the H. Some researchers suggest that multiple infection could affect the histologic changes of gastric mucosa [ 31 , 36 ]. Previous study has shown that there are differences in histological changes in the antrum and the body of the stomach between the patients with mixed H. Mixed infection was associated with significant higher rate of presence of intestinal metaplasia in the antrum [ 31 ].
In another study, mixed H. Recently, since clarithromycin has become widely used for various indications including respiratory infections or nontuberculous mycobacterial infections, the prevalence of H. Resistance to other antibiotics has also been increasing, leading to a steady decrease in eradication rates. This problem is especially a concern in local areas with high clarithromycin, quinolone, or metronidazole resistance.
Eradication rates with previously efficacious regimens have been decreasing worldwide [ 5 , 6 , 39 , 40 ]. The decreasing eradication rate and increasing resistance to antibiotics support the importance of antimicrobial susceptibility testing and the need for new efficacious eradication regimens. Given that co-infecting H. It has been suggested that heteroresistance of H.
This implies that the presence of multiple infections should be considered as a possible cause of failure in the treatment of H.
Besides, there are previous reports that show H. This means that antibiotic resistance of H. Therefore, it should be remembered that the etiology of heteroresistant antibacterial phenotypes is complex, and not only confined to infection with multiple strains. Considering these points, it might be a reasonable strategy to obtain specimen from different gastric sites for H. These situations may include when multiple infection rate is not negligible and high antibiotic resistance is anticipated, for example, in areas with both high H.
This strategy may also be useful when rescue therapy is considered. There are some limitations in this study. First, only one type of primer was used. Although H. However, antimicrobial susceptibility testing and the analysis of virulence factors supported the robustness of our RAPD fingerprinting results in identifying different H. Second, clinical records of subsequent eradication therapy and results in the patients were not available.
Third, as the number of patients included was small, and the patients with single H. According to the enrollment criteria in our study, we only included patients with successful H. So this study does not give information comparing the clinical characteristics of H. As the cases eventually showing only one positive culture in either antrum or body were not enrolled in this study, the true prevalence of multiple infection among the whole population with H. In conclusion, the presence of two H.
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Gastrointest Endosc Clin N Am — J Infect Dis e7. World J Gastroenterol 18 11 — Download references. The authors thank the members of oncology diagnostic unit, Faculty of Medicine, Suez Canal University. Mohamed A. Mohamed, Mohamed M. You can also search for this author in PubMed Google Scholar. LT contributed to the study design, processing of tissue, and nucleic acid extraction and amplification; analyzed and interpreted the patient data regarding sequencing results and blast analysis; and is a major contributor in writing the manuscript.
MF contributed to the endoscopy-guided tissue collection and collection and statistical analysis of patient data; interpreted the data; and is a minor contributor in writing the manuscript. MR contributed in the study design and is a major contributor in writing the manuscript. TS performed the histological examination of the gastric tissues.
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