Methods to monitor autophagy in H. pylori vacuolating cytotoxin A  (VacA)-treated cells.
Helicobacter pylori is a gram negative pathogen that infects at least half of the world's population and is associated not only with gastric cancer but also with other diseases such as gastritis and peptic ulcers. Indeed, H. pylori is considered the single most important risk factor for the development of gastric cancer. The vacuolating cytotoxin VacA, secreted by H. pylori, promotes intracellular survival of the bacterium and modulates host immune responses. In a recent study, we reported that VacA induces autophagy. Multilamellar autophagosomes are detected in gastric epithelial cells that are distinct from the large vacuoles formed by VacA. Furthermore, inhibition of autophagy stabilizes VacA and reduces vacuolation in the cells indicating that the toxin is being degraded by autophagy, thus limiting toxin-induced host cell damage. Many of the methods that were used for this study are commonly employed techniques that were adapted for H. pylori infection and VacA intoxication. In this paper, we describe the various methods and specific protocols used for the assessment and monitoring of autophagy during H. pylori infection.
Raju
Deepa
D
Cell Biology Program, Hospital for Sick Children, Toronto, ON, Canada.
Jones
Nicola L
NL
D000818:Animals; D001343:Autophagy / Q000187:drug effects*; D001426:Bacterial Proteins / Q000494:pharmacology*; D001431:Bacteriological Techniques; D002477:Cells / Q000187:drug effects* / Q000382:microbiology / Q000473:pathology; D002478:Cells, Cultured; D003603:Cytotoxins / Q000494:pharmacology; D019937:Diagnostic Techniques and Procedures; D005455:Fluorescent Antibody Technique / Q000379:methods; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D016481:Helicobacter Infections / Q000175:diagnosis / Q000473:pathology; D016480:Helicobacter pylori / Q000737:chemistry; D006801:Humans; D051379:Mice; D008854:Microscopy, Electron / Q000379:methods; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D003603:Cytotoxins; C477012:MAP1LC3A protein, human; D008869:Microtubule-Associated Proteins; C074689:VacA protein, Helicobacter pylori; D049452:Green Fluorescent Proteins