patchouli-alcohol and Helicobacter-Infections

Helicobacter pylori infects almost half of the world population and is listed as a type I carcinoma factor since 1994. Pogostemon cablin (Blanco) Benth. (Labiatae) has been used to treat gastro-intestinal diseases for thousands of years in many east Asian countries, and the key ingredient, patchouli alcohol (PA), has been observed to exert anti-H. pylori and anti-urease activities.. We investigated the effect of PA on H. pylori urease and its subsequent influence on macrophage phagosome maturation and function.. In H. pylori experiment, the berthelot method and pH shock assay were adopted to evaluate the effect of PA on extracellular and intracellular H. pylori urease. And then, Q-PCR and Western blot were carried out to analyze the alterations in the expression of urease-related genes and proteins after PA treatment. In the H. pylori and macrophage cell (RAW264.7) co-culture experiment, the effects of PA on H. pylori-induced phagocytosis and intracellular killing of RAW264.7 were investigated using gentamycin protection assay, and the underlying mechanism was explored by immunofluorescence.. PA at 25 and 50 μM inhibited intracellular H. pylori urease activity but not isolated urease by down-regulating the gene expression levels of ureB, ureE, ureI and nixA and reducing the protein expression level of UreB, thereby inhibiting the acid resistance of H. pylori. PA also recovered the function of macrophage bacterial digestion, and prior treatment with ammonium chloride inhibited the efficacy of PA.. PA suppressed intracellular H. pylori urease function and maturation, which increased macrophage digestion ability.

Topics: Animals; Anti-Bacterial Agents; Bacterial Proteins; Enzyme Inhibitors; Gene Expression Regulation, Bacterial; Helicobacter Infections; Helicobacter pylori; Macrophages; Mice; Phagocytosis; RAW 264.7 Cells; Sesquiterpenes; Urease

Neutrophil infiltration typically occurs in Helicobacter pylori (H. pylori)-induced acute gastritis; however, this immune response fails to eradicate H. pylori in vivo. Moreover, reactive oxygen species (ROS), which are generated by neutrophils, cause severe damage to gastric mucosa. Patchouli alcohol (PA) has been reported to have effective anti-oxidative and anti-H. pylori activities, and we investigated its effects on H. pylori-induced neutrophil recruitment and activation in this research. In neutrophil recruitment experiment, H. pylori was injected into rat air pouch to explore the effects of PA (10, 20 and 40 mg/kg) on acute inflammatory response. The results revealed that PA significantly reduced the weight of exudate and the number of neutrophils in the air pouch. Meanwhile, remarkable decrements in TNF-α and IL-8 levels in exudates were observed. In neutrophil activation experiment, rat neutrophils were isolated and activated by using 50 μg/mL H. pylori water-soluble surface protein with or without the treatment of PA (5, 10 or 20 μmol/L). Results indicated that PA not only significantly inhibited the production of ROS, but also reduced the gene and protein expressions of p22/p47-phoxes, and the binding of p22/p47-phoxes. Furthermore, the influence of PA on the neutrophil activation genes of H. pylori (h-nap and sabA) was investigated, and the results showed that expressions of h-nap and sabA were remarkably decreased after PA treatment. In conclusion, PA reduced the recruitment and activation of neutrophils induced by H. pylori, as shown by its inhibition of pro-inflammatory factor generation, p22/p47-phoxes function and H. pylori neutrophil activation-related gene expression.

Topics: Adhesins, Bacterial; Animals; Anti-Inflammatory Agents; Cytokines; Gene Expression Regulation, Bacterial; Helicobacter Infections; Helicobacter pylori; Male; NADPH Oxidases; Neutrophil Infiltration; Neutrophils; Rats, Sprague-Dawley; Reactive Oxygen Species; Sesquiterpenes

(-)-Patchouli alcohol (PA), the major active principle of Pogostemonis Herba, has been reported to have anti-Helicobacter pylori and gastroprotective effects. In the present work, we aimed to investigate the possible protective effect of PA on H. pylori urease (HPU)-injured human gastric epithelial cells (GES-1) and to elucidate the underlying mechanisms of action. Results showed that pre-treatment with PA (5.0, 10.0, 20.0μM) was able to remarkably ameliorate the cytotoxicity induced by 17.0U/mg HPU in GES-1 cells. Flow cytometric analysis on cellular apoptosis showed that pre-treatment with PA effectively attenuated GES-1 cells from the HPU-induced apoptosis. Moreover, the cytoprotective effect of PA was found to be associated with amelioration of the HPU-induced disruption of MMP, attenuating oxidative stress by decreasing contents of intracellular ROS and MDA, and increasing superoxide dismutase (SOD) and catalase (CAT) enzymatic activities. In addition, pre-treatment with PA markedly attenuated the secretion of nitric oxide (NO) and pro-inflammatory cytokines such as interleukin-2 (IL-2), interleukin-4 (IL-4) and tumor necrosis factor-α (TNF-α), whereas elevated the anti-inflammatory cytokine interleukin-13 (IL-13) in the HPU-stimulated GES-1 cells. Molecular docking assay suggested that PA engaged in the active site of urease bearing nickel ions and interacted with important residues via covalent binding, thereby restricting the active urease catalysis conformation. Our experimental findings suggest that PA could inhibit the cellular processes critically involved in the pathogenesis of H. pylori infection, and its protective effects against the HPU-induced cytotoxicity in GES-1 cells are believed to be associated with its anti-apoptotic, antioxidative, anti-inflammatory and HPU inhibitory actions.

Topics: Apoptosis; Bacterial Proteins; Catalysis; Cell Line; Cytokines; Cytoprotection; Helicobacter Infections; Helicobacter pylori; Humans; Inflammation; Intestinal Mucosa; Oxidative Stress; Pogostemon; Sesquiterpenes; Urease