Browsing by Subject "chymase"

In atherosclerosis, cholesterol accumulates in cholesterol-loaded macrophages (foam cells) forming cholesterol plaques in the arterial intima. Reverse cholesterol transport (RCT) is a mechanism in which HDL and its major structural protein apolipoprotein-A-1 (apoA-1) remove cholesterol from the foam cells and take it to the liver for its final excretion from the body in the faeces. An impaired removal of cholesterol from the foam cells is a potential contributor to a reduced RCT, which is related to a higher incidence of coronary heart disease. Chymase, a neutral protease of mast cells (MCs), is widely distributed in the connective tissue of most vertebrates and able to degrade apoA-1. After the degradation, HDL-particles are unable to interact with the ABCA-1 transporter protein on the surface of macrophages, which mediates efflux of cholesterol from the macrophage foam cells to HDL particles. It has been shown that chymase derived from rat peritoneal MCs is able to degrade apoA-1 even in the presence of blood plasma which contains natural inhibitors for chymase (α-2-macroglobulin and α-1-antichymotrypsin). In the present study we wanted to find out if mouse mast cell protease 4 (mMCP-4) isolated from peritoneal mast cells is able to maintain its enzymatic activity even in the presence of mouse serum and intraperitoneal fluid. A small molecular weight compound (S-2586) was used as a substrate. In the in vitro experiments a sonicated MC preparation that contains active chymase was used and the activity of chymase was measured in the presence of varying concentrations of plasma and intraperitoneal fluid. In the in vivo experiments we evaluate whether mast cell-dependent proteolysis of HDL particles does occur, and whether such modification inhibits their efficiency in inducing cellular cholesterol efflux in vitro. We found that both serum and intraperitoneal fluid inhibited chymase activity, serum to a higher extent. Systemic activation of MCs in mast cell-competent mice, but not in mast cell-deficient mice, in vivo led to a decreased ability of plasma and intraperitoneal fluid to act as cholesterol acceptors from cultured cholesterol-loaded macrophages. Local activation of peritoneal mast cells also blocked the cholesterol efflux-inducing effect of intraperitoneally injected human apoA-1. This work was performed at the Wihuri Research Institute. Licenses for animal work were approved by the Finnish Laboratory Animal Experiment Committee (Suomen eläinkoelautakunta, ELLA). Laboratory animals (female NMRI mice) were from the Viikki Laboratory Animal Centre of the University of Helsinki and the mast cell deficient strain of mice W-sash c-kit mutant KitW-sh/W-sh were from the Jackson Laboratory (BarHarbor, Maine). The work was supervised by the director of the Research Institute Petri Kovanen MD PhD and Miriam Lee-Rueckert PhD. Laboratory assistance was perceived from the technicians of the Wihuri Research Institute.