Iron in synovial fluid: removal by lactoferrin, and relationship to iron regulatory protein (IRP) activity.

Rheumatoid arthritis is a chronic inflammatory polyarthritis characterised by an inflammatory cell infiltration of the synovial membrane. I t has been suggested that in these patients an aggressive oxidant such as the OH’ radical may be responsible for membrane damage and other harmful effects within the synovial membrane [l]. Iron, in a form able to catalyse OH’ formation may be released from iron binding proteins such as transferrin, lactoferrin and ferritin at low pH values found in the microenviroment beneath adherent macrophages, and accumulates in the synovium and may also be present in synovial fluid. If levels of synovial iron-binding proteins could be increased, “free” iron, and hence tissue damage, might be reduced. Lactoferrin, due to its high affinity for iron even at moderately acid pH, is of particular interest in this respect. We have therefore investigated wether “free” iron in synovial fluid is related to the degree of inflammation,and whether it affects the iron status of cells i n the fluid by measuring the activity of iron-regulatory protein (IRP), which regulates transferrin receptor and ferritin post-transcriptionally by binding to their mRNA’s under low iron conditions [2]. Synovial fluids were obtained by knee aspiration from 25 patients (20 women and 5 men, age range 19-89). Of these, 21 fulfilled the criteria for rheumatoid arthritis [3], 2 had juvenile chronic arthritis, 1 psoriasis and 1 osteoarthritis. Synovial fluid samples were centrifuged to sediment the cells. The supernatant was used to determine “free” iron by the bleomycin assay [4] and the cells to determine iron status by measuring total and active IRP by mobility shift assay in the presence or absence of 2-mercaptoethanol respectively [5]. Synovial fluids known to contain “free” iron were used to test the binding activity of lactoferrin by incubating them overnight at 4°C with lactoferrin (human or mouse) or transferrin, and then determining “free” iron by the bleomycin assay.