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Annals of the Rheumatic Diseases logoLink to Annals of the Rheumatic Diseases
. 1998 May;57(5):309–314. doi: 10.1136/ard.57.5.309

Iron, lactoferrin and iron regulatory protein activity in the synovium; relative importance of iron loading and the inflammatory response

C Guillen 1, I McInnes 1, H Kruger 1, J Brock 1
PMCID: PMC1752600  PMID: 9741316

Abstract

OBJECTIVES—To determine the ability of lactoferrin in rheumatoid arthritis (RA) synovial fluid to bind "free" iron, and to study the regulatory mechanisms therein that control iron homeostasis.
METHODS—"Free" iron was determined by the bleomycin assay and lactoferrin concentrations by enzyme linked immunosorbent assay. The activities of iron regulatory protein (IRP) and NF-κB in synovial fluid cells were assayed by mobility shift assay.
RESULTS—30% of synovial fluids contained "free" iron and in these, lactoferrin concentrations were significantly lower than in those with no "free" iron (p<0.01). Addition of exogenous lactoferrin consistently reduced the amount of "free" iron in positive synovial fluids. IRP activity in synovial cells did not correlate with synovial fluid iron concentrations but did correlate with NF-κB activation and with serum C reactive protein.
CONCLUSION—Lactoferrin may prevent iron mediated tissue damage in RA by reducing "free" synovial iron concentration when inflammatory stimuli have disregulated IRP mediated iron homeostasis.

 Keywords: lactoferrin; rheumatoid arthritis; inflammation

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Figure 1  .

Figure 1  

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"Free" iron in synovial fluid of RA patients. The results represent the concentrations of "free" iron in RA patients using the bleomycin assay at pH 5.3. 

Figure 2  .

Figure 2  

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Lactoferrin (A) and transferrin (B) in synovial fluid from RA patients. The results correspond to the synovial fluid lactoferrin and transferrin concentrations in RA patients segregated for the presence or absence of "free" iron. Lactoferrin concentrations were significantly lower (p<0.01) in fluids containing "free" iron. There was no significant difference in transferrin concentrations.

Figure 3  .

Figure 3  

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Binding of iron in synovial fluid by exogenous lactoferrin and transferrin. Synovial fluid samples were incubated overnight at 4°C with different concentrations of lactoferrin or transferrin and "free" iron was measured by the bleomycin assay. The results show the percentage decrease in "free" iron (mean (SEM); n=7) for each concentration of exogenous lactoferrin or transferrin.

Figure 4  .

Figure 4  

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Correlation between iron regulatory protein (IRP) activity in synovial fluid cells of RA patients with "free" iron (A) and CRP (B). IRP activity was calculated from the ratio of intensity of IRE-IRP complexes in mobility shift assays performed with or without 2-mercaptoethanol. Serum CRP concentrations correlated with IRP activity (p=0.013)

Figure 5  .

Figure 5  

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Mobility shift assay of IRP (A) and NFκB (B) from synovial fluid mononuclear and polymorphonuclear cells from a representative case of three RA patients. In panel A, lanes 1 and 2 correspond to active IRP in synovial fluid mononuclear cells and lanes 3 and 4 to active IRP in synovial fluid polymorphonuclear cells. Samples in lanes 1 and 3 were treated with 2-mercaptoethanol (2-ME) to convert inactive IRP to the active form (see methods). In panel B, lane 1 corresponds to NFκB activity in synovial fluid mononuclear cells and lane 2 to NFκB activity in synovial fluid polymorphonuclear cells.

Selected References

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