Macrophage migration inhibitory factor is increased in the urine of patients with urinary tract infection: Macrophage migration inhibitory factor-protein complexes in human urine

Purpose: MIF is a proinflammatory cytokine present in preformed stores in human urothelium. In animal Models of bladder inflammation, including bacterial cystitis, MIF is up-regulated in the bladder and released from the bladder as a high molecular weight complex. We compared urine MIF amounts in patients with UTI to that in patients without UTI, and we examined and identified MIF-protein complexes in urine. Materials and Methods: Using enzyme-linked immunosorbent assay we compared MIF levels in the urine of 14 patients with UTI to levels in 16 controls with no UTI. Western blotting under native, denaturing and reducing conditions was done to examine MIF complexes found in urine. Mass spectrometry identified MIF associated proteins in urine, while coimmunoprecipitation confirmed the associations. Results: Mean urine MIF amounts SEM determined by enzyme-linked immunosorbent assay were significantly greater in 14 patients with UTI compared to that in 16 controls (1.96 +/- 0.40 vs 0.59 +/- 0.09 ng/mg creatinine, p < 0.01). Western blotting under denaturing conditions showed several high molecular weight complexes (100 to 165 kDa) that increased in UTI urine as well as typical, monomeric MIF (12 kDa). Mass spectrometry identified associated MIF proteins, including ceruloplasmin, albumin and uromodulin. Co-immunoprecipitation confirmed mass spectrometry findings and also identified MIF interaction with alpha-2-macroglobulin. Conclusions: Increased urine MIF amounts in patients with bacterial cystitis support our experimental evidence showing a role for MIF in pelvic visceral inflammation. The novel finding of an association of MIF with other urine proteins suggest that the physiologically relevant form of MIF may be an MIF-protein complex.