Date of Award

5-2012

Document Type

Thesis

Degree Name

Master of Dental Science (MDS)

Program

Periodontology

Research Advisor

Jegdish P. Babu, M.S., Ph.D.

Committee

Anastasios Karydis, D.D.S., M.S., Ph.D. Sidney H. Stein, D.M.D., M.S., Ph.D. David A. Tipton, D.D.S., Ph.D.

Abstract

The cranberry (species: Vaccinium macrocarpon, family: Ericaceae) is a polyphenolicrich berry fruit native to North America and exhibits numerous beneficial properties for human health, such as inhibition of human cancer cell line proliferation and prevention of urinary tract colonization by bacterial pathogens. The proanthocyanidin rich cranberry fraction has long been speculated to possess the capacity to reduce the production of inflammatory mediators by LPSstimulated macrophages and gingival fibroblasts and to inhibit host extracellular matrix destructive enzyme production and activity.

The purpose of this investigation was to examine the effect of cranberry non dialyzable material (NDM), which was shown to be rich in proanthocyanidins; this has a suppressive effect on production of inflammatory cytokines mediated by LPS which are known to play a major destructive role in periodontal disease. In addition to the cytokines, stimulation of superoxide ions by the monocytes also contributes to the severity of periodontal disease. In this study we chose to investigate the effect of NDM on LPS-mediated stimulation of the cultured monocytic cells (THP-1) to secrete the two cytokines; IL-1β and TNF-α, and also on superoxide release. We chose to study the effect of NDM on biological functions of isolated LPS from periodontal pathogens. LPS was isolated from P. gingivalis, T. denticola and F. nucleatum.

Secretion of the two cytokines by the cultured monocytes was found to be dosedependent upon the concentration of LPS. Secretion of TNF-α varied from 110±28 to 610±42 picograms/ml (pg/ml) when stimulated cells with 1.0 and 20 ng/ml of LPS. Similarly, IL-1β secretion varied from 49±6 to 196±16 pg/ml. The LPS of both T. denticola and F. nucleatum also exhibited similar phenomenon. In the presence of NDM (20 µg/ml), the cells secreted 80- 90% less of the two cytokines when stimulated with LPS. LPS isolated from the same three oral pathogens grown in the presence of NDM (10-20 µg/ml) also failed to stimulate the monocytes compared to the LPS isolated from bacteria grown in culture media alone. Interestingly, when the THP-1 cells were first incubated with NDM, the cranberry components also caused a significant decrease in the secretion of cytokines. The results of the study strongly indicate the suppressive effect of NDM on LPS-mediated secretion of cytokines by the monocytic cells. It may be possible that the NDM may be interfering with LPS interaction with LPS binding protein and the CD-14, a known receptor on the monocytes which triggers the secretion of cytokine molecules.

The LPS of pathogenic bacteria induces the monocytic cells to secrete superoxide ions; a mechanism which aids in tissue destruction. Our results showed that the LPS isolated from all three pathogens stimulated cells to secrete superoxide ions several folds higher (65 to 89%) than the control cells which were not exposed to LPS. Cultured monocytic cells when incubated with LPS along with NDM secreted significantly lesser superoxide ions (60-70% reduction). The results of the study clearly demonstrated that the cranberry NDM alters the biological functions of bacterial LPS. The NDM inhibited the secretion of cytokines and also superoxide ions by the cultured monocytes in response to LPS.

Therapeutic agents that modulate host inflammatory mediators have shown promise for managing adult periodontitis and may be highly useful for individuals with a substantially increased risk for periodontitis. In addition to the previously recognized inhibitory effect of the cranberry non-dialyzable material fraction on the aggregation of oral bacteria and dental biofilm formation we showed that this fraction was a potent inhibitor of the pro-inflammatory cytokines and superoxide ions induced by LPS of periodontopathogens. This provides promising perspectives for the development of novel host-modulating therapies for adjunctive treatments of periodontitis or other inflammatory diseases by use of the high molecular weight constituents from cranberries.

DOI

10.21007/etd.cghs.2012.0276

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