Abstract

Quorum-sensing molecules regulate the behavior of bacteria within biofilms and at the same time elicit an immune response in host tissues. Our aim was to investigate the regulatory role of dihydroxy-2,3-pentanedione (DPD), the precursor of universal autoinducer-2 (AI-2), and its analogs (ethyl-DPD, butyl-DPD and isobutyl-DPD) in the integrity of gingival epithelial cells. Human gingival keratinocytes were incubated with four concentrations (10μmol L-1 , 1μmol L-1 , 100nmol L-1 and 10nmol L-1 ) of DPD and its analogs for 24hours. The numbers of viable cells were determined using a proliferation kit, matrix metalloproteinase (MMP)-2 and -9 activities were determined by gelatin zymography, and expression of occludin protein and occludin mRNA were determined by western blotting and RT-qPCR, respectively. Increased cell proliferation was observed in gingival keratinocytes incubated with 100nmol L-1 of butyl-DPD. MMP-9 activity was elevated in cells incubated with 10μmol L-1 of ethyl-DPD. On the other hand, MMP-2 activity did not show any significant change when gingival keratinocytes were incubated with or without DPD or analogs. Western blot analyses demonstrated five forms (105, 61, 52.2, 44 and 37kDa) of occludin. Incubation with 1μmol L-1 and 100nmol L-1 of DPD and with 10nmol L-1 of ethyl-DPD increased dimeric (105kDa) forms of occludin, while incubation with 100nmol L-1 of isobutyl-DPD increased monomeric (61kDa) forms. DPD and ethyl-DPD decreased, and 100nmol L-1 of isobutyl-DPD and 10nmol L-1 of butyl-DPD increased, the monomeric (52.2kDa and 44kDa) forms of occludin, whereas ethyl-DPD decreased and isobutyl-DPD increased, the low-molecular-weight (37kDa) forms. According to RT-qPCR analysis, the exposure of gingival keratinocytes to 10μmol L-1 of isobutyl-DPD up-regulated expression of occludin. The results indicate that isobutyl-DPD has the potential to enhance the integrity of the epithelium by stimulating the formation of occluding, without affecting the proliferation or gelatinolytic enzyme activities of the exposed cells. The modulatory effect of an AI-2 analog on the epithelial cell response is shown for the first time.

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