Platelet‐Activating Factor Mediates Helicobacter pylori Lipopolysaccharide Interference with Gastric Mucin Synthesis

Abstract

SummaryPlatelet-activating factor (PAF) is now recognized as the most proximal mediator of cellular events triggered by bacterial infection. In this study, we report that a specific PAF antagonist, BN52020, impedes the reduction in mucin synthesis evoked in gastric mucosal cells by H. pylori LPS. The impedance by BN52020 of the LPS inhibitory effect on mucin synthesis was blocked by wortmannin, an inhibitor of phosphatidylinositol 3-kinase (PI3K), which also obviated the inhibitory effect of BN52020 on the LPS-induced upregulation in apoptosis, TNF-a, and NO generation. A reduction in the impedance by BN52020 of the LPS detrimental effect on mucin synthesis was also attained with cNOS inhibitor, L-NNA, whereas NOS-2 inhibitor, 1400W caused a potentiation in the impedance effect of BN52020. However, while 1400W and BN52020 countered the potentiating effect of wortmannin on the LPS-induced decrease in mucin synthesis, a further exacerbation of the potentiating effect of wortmannin was attained in the presence of cNOS inhibitor, L-NNA. Our findings suggest that PAF, through the interference with PI3K-dependent cNOS activation, plays a critical role in influencing the extent of pathological consequences of H. pylori infection on the synthesis of gastric mucin.

Experimental study on gastroprotective efficacy and mechanisms of luteolin-7-O-glucoside isolated from Ophiorrhiza mungos Linn. in different experimental models

“…Arginase level was measured and the results are expressed as U/mg proteins (del Ara et al, 2002). In order to determine the levels of superoxide dismutase (SOD), and apoptosis, the methods of Sun, Oberley, and Li (1988), and Slomiany and Slomiany (2004) were used, respectively, and their results are expressed as U/mg protein.…”

Role of constitutive nitric oxide synthase S-nitrosylation in Helicobacter pylori-induced gastric mucosal cell apoptosis: effect of ghrelin

“…In the experiments evaluating the effect of ghrelin (rat, Sigma), cNOS inhibitor, L-NAME, iNOS inhibitor, 1400W, Src inhibitor, PP2, Akt inhibitor, SH-5 (Calbiochem), and ascorbate (Sigma), the cells were first preincubated for 30 min with the indicated dose of the agent or vehicle before the addition of the LPS. The viability of cell preparations before and during the experimentation, assessed by Trypan blue dye exclusion assay (Slomiany and Slomiany 2004), was greater than 97%.…”

“…Following centrifugation, the supernatant containing the cytoplasmic histone-associated DNA fragments was diluted and reacted with immobilized antihistone antibody. After washing, the retained complex was reacted with anti-DNA peroxidase, and probed with ABTS reagent for spectrophotometric quantification (Slomiany and Slomiany 2004). To assess NO production in the mucosal cells, we measured the stable NO metabolite, nitrite, accumulation in the culture medium using Griess reaction (Wagner et al 1982).…”

“…A 100 ll aliquot of spent culture medium was incubated for 10 min with 0.1 ml of Griess reagent and the absorbance was measured at 570 nm. The concentration of nitrite, expressed as lM/ml, was calculated with sodium nitrite as standard (Slomiany and Slomiany 2004).…”

Cytosolic phospholipase A 2 activation in Helicobacter pylori lipopolysaccharide-induced interference with gastric mucin synthesis

“…Aliquots of gastric mucosal cell suspension (1 ml) were transferred to DMEM in culture dishes containing [ 3 H]glucosamine (110 mCi), and incubated according to previously described conditions in absence or the presence of H. pylori LPS at 400 ng/ml (25). To study the effect of cPLA 2 inhibitor, MAFP, PAF inhibitor, BN52020, ERK1/2 inhibitor, PD98059, and PI3K inhibitor, wortmannin, the cells were preincubated for 30 min with the indicated dose of the agent or the vehicle before the addition of the LPS.…”