Histatin 5 attenuates IL-8 dendritic cell response to gingivalis Hemagglutinin B

Borgwardt DS
Source: Other
Publication Date: (2011)
Issue: 0:0: Thesis
Research Area:
Immunotherapy / Hematology
Basic Research
Cells used in publication:
Dendritic cell (NHDC), human
Species: human
Tissue Origin: blood
Histatins, a group of proteins produced by human salivary glands, have a variety of innate immune functions including the ability to: kill oral microorganisms, neutralize toxins, inactivate protease/collagenase activities, inhibit co-aggregation of oral bacteria, and inhibit lipopolysaccharide mediated activities. Hemagglutinin B (HagB), a virulence factor of the periodontal pathogen Porphyromonas gingivalis, induces a robust cytokine and chemokine response in human myeloid dendritic cells. In this study, I hypothesize that histatin 5 can attenuate a HagB-induced chemokine response. Objectives: To characterize an expanded cytokine and chemokine response induced in human myeloid dendritic cells by HagB, and to determine if prior incubation of HagB with histatin 5 attenuates these responses. Methods: In my first experiment, 0.040 ?M HagB was mixed with dilutions of histatin 5 and histatin 8 (Sigma, 0.04 to 40.0 ?M), incubated at 37?C for 30 minutes, and added to 2 x 104 human myeloid dendritic cells (Lonza, Walkersville, MD). At 24 hours, culture media was removed, and 6 cytokines and chemokines (pg/ml) were determined in cell-free supernatants (Millipore, Billerica, MA) using the Luminex 100 IS instrument (Luminex, Austin, TX). In my second experiment, 0.040 ?M HagB was mixed with 40.0 ?M histatin 5 only (e.g., 1:1000), incubated at 37?C for 30 minutes, and added to 2 x 104 human myeloid dendritic cells. At 0, 1, 2, 4, 8, 16, and 24 hours post-inoculation, culture media was removed, and 26 cytokines and chemokines (pg/ml) were determined in cell-free supernatants. Results: In both experiments, human myeloid dendritic cells incubated with HagB produced Th1, Th2, and Th17 cytokines (IL-2, IL-12(p70), IFN-?, IL-3, IL-4, IL-5, , IL-15, IL-17); pro-inflammatory cytokines (IL-1?, IL-1?, IL-6, TNF-?, IL-12(p40); anti-inflammatory v cytokines (IL-10, IL-13, IFN?2); chemokines (CXCL8/IL-8, CXCL10/IP-10, CCL2/MCP-1, CCL3/MIP-1?, MIP-1b, CCL11/eotaxin); and colony stimulating factors (IL-7, G-CSF, GM-CSF). Histatin 5 significantly attenuated (p < 0.05) the IL-8 response induced by HagB at 8 - 16 hours and to a lesser extent, the IL-6, GM-CSF, MCP-1, MIP-1a, MIP-1ß, and TNF-a response. Conclusion: Histatin 5 is an important salivary component capable of attenuating an IL-8 response. Together with human beta defensin 3, another peptide previously shown to attenuate pro-inflammatory cytokines, histatin 5 may help control and contain oral infection and inflammation by down regulating IL-8 chemotactic response.