The role of chondroitin sulfate chains of urinary trypsin inhibitor in inhibition of LPS-induced increase of cytosolic free Ca2+ in HL60 cells and HUVEC cells.

Preincubation of HL60 cells and HUVEC cells with urinary trypsin inhibitor (UTI) inhibited increase of cytosolic free Ca2+ induced by LPS. In contrast, an increase of cytosolic free Ca2+ induced by LPS was not inhibited by deglycosylated UTI, UTI treated with monoclonal antibody of chondroitin sulfate. 45Ca2+ binding showed that UTI binds 45Ca2+ dose-dependently. Scatchard plot analysis showed that UTI has two binding sites for Ca2+, a high affinity binding site (Kd=15 microM) and a low affinity site (Kd=150 microM), and that UTI has more than 70 Ca2+ binding sites per molecule. The Ca2+ binding capacity of deglycosylated UTI and UTI treated with monoclonal antibody of chondroitin sulfate was markedly depressed. Furthermore, UTI forms multi-polymers in the presence of Ca2+ as demonstrated by gel filtration and agarose gel electrophoresis. These results suggest that UTI is a physiological Ca2+ chelator on the cells and that the action is due to chondroitin sulfate chains of UTI. Copyright 1997 Academic Press.