Complexes of tissue kallikrein with protein C inhibitor in human semen and urine.
An ELISA was developed for quantifying the complex between tissue kallikrein (tKK) and protein C inhibitor (PCI) (tKK:PCI) in seminal plasma and urine. The ELISA used purified tKK:PCI complex as a standard and was specific for this complex with a detection limit of about 1.1 pM. Purified tKK:PCI complex was obtained from human urine and was 95% homogeneous as judged by SDS/PAGE. The 90-kDa band corresponding to the purified tKK:PCI complex reacted with anti-tKK and anti-PCI antibodies as judged by immunoblotting. Seminal plasma collected in the absence of extrinsic inhibitors contained 1.8 +/- 0.6 nM tKK:PCI complex and 4.7 +/- 2.8 nM immunoreactive tKK (mean +/- SD, n = 10), which indicates that about 28% of the total tKK immunoreactivity is forming complexes with PCI. When semen was collected in the presence of tKK inhibitors it had only about 6% of the tKK complexed to PCI. In vitro studies showed that the tKK:PCI complex formation in semen was accomplished in about 1 h and that heparin stimulated both the rate and the extent of complexation of tKK with PCI. Native urine showed low levels of tKK:PCI complex, but after dialysis urine had 0.17 +/- 0.05 nM complex. Formation of tKK:PCI complex in urine and semen was also demonstrated by immunoblotting. These results suggest that PCI is a physiological inhibitor of tKK and provide additional evidence of the involvement of PCI in human reproduction.
Research Center, La Fe University Hospital, valencia, Spain.
F
F
España
E
E
Fink
J
J
Sanchez-Cuenca
J
J
Gilabert
A
A
Estellés
K
K
Witzgall
D000328:Adult; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007610:Kallikreins / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D008297:Male; D008875:Middle Aged; D018046:Protein C Inhibitor / Q000378:metabolism*; D012661:Semen / Q000378:metabolism*; D020840:Tissue Kallikreins; D014568:Urokinase-Type Plasminogen Activator / Q000378:metabolism
D018046:Protein C Inhibitor; D007610:Kallikreins; D020840:Tissue Kallikreins; D014568:Urokinase-Type Plasminogen Activator