• Salminen, Aino

Abstract

Matrix metalloproteinases (MMPs) are enzymes that are responsible for the degradation of the extracellular matrix (ECM) during development, repair, and remodeling of tissues. MMP-8, also known as neutrophil collagenase, is released from neutrophils when they enter the tissues at the site of inflammation. An imbalance in MMP-8 activity leads to excess degradation of tissues and destructive inflammation, such as periodontitis. Elevated concentrations of MMP-8 in serum and plasma are also associated with cardiovascular diseases (CVDs). Doxycycline has inhibitory effects against MMPs in addition to its well-known property of being antimicrobial. This thesis investigated the genetics of MMP-8, the effect of MMP-8 and its inhibitors on lipid metabolism, and the potential of salivary MMP-8 in diagnostics of periodontitis. We performed a genome-wide association study (GWAS) in two independent populations with a total of 6049 individuals to identify genetic variants and molecular mechanisms that affect serum MMP-8 concentrations. In addition, we studied whether MMP-8-associated genetic variants are related to increased risk of CVDs in over 20 000 individuals. We discovered that genetic polymorphism in the gene of complement factor H (CFH) is strongly associated with the concentrations of MMP-8 in serum. By conducting functional experiments with isolated neutrophils, we found that genetic variation of CFH affected the release of MMP-8 from neutrophils in response to complement activation. In addition, genetic polymorphism in the locus containing the genes of S100 calcium binding proteins A8, A9, and A12 was associated with serum and plasma MMP-8 levels and also with the prevalence and incidence of CVDs. We studied the effect of MMP-8 and its inhibitors on lipid metabolism by conducting cell experiments, in an MMP-8-knockout mouse model, and in a placebo-controlled clinical trial of two years. We discovered that MMP-8 cleaved apolipoprotein A-I (apoA-I), the main protein component of HDL. MMP-8 significantly reduced the ability of HDL to promote cholesterol efflux from cholesterol-loaded macrophages. The cleavage of apoA-I by MMP-8 and the reduction in its cholesterol efflux capacity was inhibited by doxycycline at clinically attainable doses. MMP-8 deficient mice had significantly lower serum triglyceride levels and larger HDL particle size compared to wild type mice. In the clinical trial, the subjects treated with subantimicrobial-dose doxycycline (SDD) displayed a significant increase in cholesterol efflux from macrophages to the serum compared to the baseline, whereas the efflux levels did not change in the placebo group over the study period. We studied the association of three salivary biomarkers, MMP-8, interleukin-1β, and Porphyromonas gingivalis, with periodontal status in 463 subjects. The salivary concentrations of MMP-8, interleukin-1β, and P. gingivalis were associated with the number of deepened periodontal pockets and the extent of alveolar bone loss. The combination of the biomarkers was more strongly associated with moderate to severe periodontitis than any of the biomarkers alone. Our results indicate that activation of the complement system, especially the alternative pathway, contributes significantly to the concentrations of MMP-8 in serum. Genetic polymorphism in S100A8/A9/A12 locus affects circulating MMP-8 levels, and is associated with CVDs. These results emphasize the role of inflammation and the immune system in CVDs. Proteolysis of apoA-I by MMP-8 may disturb HDL metabolism and reverse cholesterol transport, which leads to accumulation of cholesterol in the vessel walls and accelerated atherosclerosis. Inhibition of MMP-8 by doxycycline may reduce the risk of CVDs, especially in vulnerable individuals such as periodontitis patients. Saliva MMP-8, particularly when combined with other biomarkers, has great potential in the diagnostics of periodontitis. MMP-8 in saliva reflects the health of the oral cavity, whereas circulating MMP-8 is associated with systemic diseases such as CVDs. Our results suggest that MMP-8 functions as a link between inflammatory disorders, such as periodontitis, and cardiovascular disorders.

Topics

• impedance spectroscopy
• experiment
• activation
• Calcium
• chemical vapor deposition