Caspase‐1‐dependent mechanism mediating the harmful impacts of the quorum‐sensing molecule N‐(3‐oxo‐dodecanoyl)‐l‐homoserine lactone on the intestinal cells

Abstract

N-(3-oxododecanoyl)-L-homoserine lactone (3-oxo-C12-HSL), a quorum-sensing (QS) molecule produced by Gram-negative bacteria in the gastrointestinal tract, adversly impacts host cells. Our previous study demonstrated that 3-oxo-C12-HSL induced a decrease in cell viability via cell apoptosis and eventually disrupted mucin synthesis from LS174T goblet cells. However, the molecular mechanism underlying cell apoptosis and whether pyroptosis was involved in this process are still unknown. In this study, we emphasized on the caspases signal pathway and sterile inflammation to reveal the harmful effects of 3-oxo-C12-HSL on LS174T goblet cells. Our data showed that 3-oxo-C12-HSL is a major inducer of oxidative stress indicated by a high level of intracellular reactive oxygen species (ROS). However, TQ416, an inhibitor of paraoxonase 2, can effectively block oxidative stress. A higher ROS level is the trigger for activating the caspase-1 and 3 cascade signal pathways. Blockade of ROS synthesis and caspase-1 and 3 cascades can obviously rescue the viability of LS174T cells after 3-oxo-C12-HSL treatment. We also found that paralleled with a higher level of ROS and caspases activation, an abnormal expression of proinflammatory cytokines was induced by 3-oxo-C12-HSL treatment; however, the blockage of TLRs-NF-κB pathway cannot restore cell viability and secretary function. These data collectively indicate that 3-oxo-C12-HSL exposure induces damages to cell viability and secretary function of LS174T goblet cells, which is mediated by oxidative stress, cell apoptosis, and sterile inflammation. Overall, the data in this study will provide a better understanding of the harmful impacts of some QS molecules on host cells and their underlying mechanism.

Original low birth weight deteriorates the hindgut epithelial barrier function in pigs at the growing stage

“…Meanwhile, the decreased SOD and GPX enzyme activity suggest that LBW limited the antioxidant capacity of the pig's hindgut mucosa. It is well known that caspase-3 is an executioner of cell apoptosis (38,39). We observed an increase in the caspase-3 gene expression and activity in the hindgut mucosa of LBW pigs.…”

Icariin and its phosphorylated derivatives alleviate intestinal epithelial barrier disruption caused by enterotoxigenic Escherichia coli through modulate p38 MAPK in vivo and in vitro

“…25 In brief, after dilution to a final concentration of 10 μM with serum-free DMEM, DCFH-DA was added to the cells after the culture medium was removed and incubated for 30 minutes at 37°C. 25 In brief, after dilution to a final concentration of 10 μM with serum-free DMEM, DCFH-DA was added to the cells after the culture medium was removed and incubated for 30 minutes at 37°C.…”

“…Intracellular ROS in IPEC-J2 cells were measured with 2′,7′-dichlorofluorescein diacetate (DCFH-DA; Sigma) as a previously reported method. 25 In brief, after dilution to a final concentration of 10 μM with serum-free DMEM, DCFH-DA was added to the cells after the culture medium was removed and incubated for 30 minutes at 37°C. Next, the cells were washed 3 times with phosphate-buffered saline (PBS).…”

The protective effect of icariin and phosphorylated icariin against LPS-induced intestinal goblet cell dysfunction

“…Cell viability was tested as described previously. 26 Briefly, cells ($2 Â 10 5 cells) were cultured in 96-well cell culture plates with treatment. After 24 h, 10 ll of the CCK-8 assay solution was added to each well and incubated for another 1 h. The optical densities were then read on a microplate reader (Molecular Devices, Sunnyvale, CA, USA) at 450 nm.…”

“…Intracellular reactive oxygen species (ROS) in LS174T cells were measured with 2 0 ,7 0 -dichlorofluorescein diacetate (DCFH-DA; Sigma) as previously reported. 26 In brief, after dilution to a final concentration of 10 lM with serum-free DMEM, DCFH-DA was added to the cells after the culture medium was removed and incubated for 30 min at 37 C. Next, the cells were washed three times with PBS. The cells were re-suspended in PBS, and the fluorescence intensity was measured for more than 10,000 cells of each sample using a FACSVerse flow cytometer (excitation/emission wavelength ¼ 504/529 nm).…”