Matrix Metalloproteinase‐9 Release from Porphyromonas gingivalis Lipopolysaccharide (LPS)‐treated Rat Brain Microglia In Vitro

Abstract

BackgroundThe putative association between periodontal disease (PD), a chronic inflammatory disease and neuroinflammation, a condition which involves brain microglia, remains under investigation. We have reported that Porphyromonas gingivalis (Pg) lipopolysaccharide (LPS), a bacterium implicated in periodontitis, activates both classical (M1‐type) and alternative (M2‐type) activation of neonatal rat microglia (BMG) with concomitant release of superoxide anion, thromboxane B2, cytokines, and chemokines (Faseb Journal 32.1 Supplement: 702.2, 2018). We hypothesized that matrix metalloproteinase 9 (MMP‐9) release by Pg‐treated microglia might be detectable by both ELISA and visualized by confocal fluorescence imaging.MethodsP. gingivalis LPS (Pg LPS) 105 ng/mL from InvivoGen (San Diego, CA) and Escherichia coli LPS (Ec LPS) 026:B6 1 ng/mL from Difco Lab, Detroit, MI (positive control) were used to treat rat BMG for 18 hours at 35.9°C in vitro. MMP‐9 release was assessed with a rat MMP‐9 ELISA kit (R&D Systems). Confocal fluorescence imaging with a Nikon A1R laser confocal microscope was used to identify BMGs using a primary mouse anti‐rat CD11b/c antibody (Ab) (AbD SeroTec, Raleigh, NC), and secondary donkey anti‐mouse Ab (Thermo Fischer, Waltman, MA). MMP‐9 was visualized with a primary rabbit anti‐rat MMP‐9 polyclonal Ab, and secondary goat anti‐rabbit Ab, both from Abcam, Cambridge, MA.ResultsMMP‐9 presence in tissue culture supernates was detectable by ELISA in both Ec LPS and Pg LPS‐treated BMG. In contrast, MMP‐9 was observed by confocal fluorescence imaging only within Ec LPS‐treated BMG but not in Pg LPS‐treated BMG.ConclusionsBoth ELISA and confocal fluorescence imaging provide additional support for our working hypothesis, because Pg LPS stimulated BMGs to release MMP‐9. Thus, our current observations extend our studies on the effect of Pg LPS on rat brain microglia activation and contribute to further characterize the putative role of Pg LPS in both periodontitis, neuroinflammation and neurodegeneration.Support or Funding InformationSupport by College of Dental Medicine, College of Graduate Studies, and Chicago College of Osteopathic Medicine, Midwestern University are gratefully acknowledged.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.