Synthesis of tissue factor pathway inhibitor in human synovial cells and chondrocytes makes joints the predilected site of bleeding in haemophiliacs.
The synthesis of tissue factor pathway inhibitor (TFPI) was investigated in cloned human synovial cells and human chondrocytes. TFPI-specific DNA transcription products of these cells were isolated, and a full-length cDNA of about 1000 base pairs was amplified by reverse transcription and polymerase chain reaction. The amplified DNA was cloned into the vector pUC 18. The TFPI coding sequence was confirmed by double stranded sequencing and was identical with that previously published for human TFPI coding nucleotide sequence from human placental cDNA (1). The inhibitory activity of TFPI in the cell medium of cultivated human chondrocytes and cloned human synovial cells was determined by a specific chromogenic substrate assay of factor Xa activity. The inhibitory activity of TFPI in the medium of human chondrocytes and cloned human synovial cells was 630-720 mU/10(8) cells and 1080-1665 mU/10(8) cells, respectively. In addition, TFPI activity in cell culture media of human chondrocytes and cloned human synovial cell was suppressed by a polyclonal goat anti-TFPI antibody directed against the inhibitory domain I and domain II. In the chromogenic substrate assay, the anti-TFPI antibody completely suppressed the inhibitory activity of TFPI in the samples.
Brinkmann
T
T
Institut für Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum Nordrhein-Westfalen, Universitätsklinik der Ruhr-Universität Bochum, Bad Oeynhausen, Germany.
Kähnert
H
H
Prohaska
W
W
Nordfang
O
O
Kleesiek
K
K
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology / Q000494:pharmacology; D000918:Antibody Specificity; D001483:Base Sequence; D002356:Cartilage / Q000166:cytology / Q000378:metabolism*; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D005167:Factor VII / Q000037:antagonists & inhibitors; D006467:Hemophilia A / Q000378:metabolism* / Q000503:physiopathology; D006470:Hemorrhage / Q000209:etiology; D006801:Humans; D008074:Lipoproteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D008969:Molecular Sequence Data; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D013583:Synovial Membrane / Q000166:cytology / Q000378:metabolism*; D013925:Thromboplastin / Q000037:antagonists & inhibitors; D014158:Transcription, Genetic
D000906:Antibodies; D018076:DNA, Complementary; D008074:Lipoproteins; C051928:lipoprotein-associated coagulation inhibitor; D012313:RNA; D005167:Factor VII; D013925:Thromboplastin