Inhibitory effect of protease inhibitor on endothelial cell activation.
BACKGROUND
Endothelial cells (EC) are important for regulating the hemostatic balance of prothrombotic and antithrombotic activities. Proinflammatory cytokines such as tumor necrosis factor-alpha (TNFalpha) play an important role in the regulation of EC and also regulate the production of plasminogen activator inhibitor type 1 (PAI-1), which is an EC-producing factor with the inhibitory activity of fibrinolysis, and the expression of intercellular adhesion molecule-1 (ICAM-1), which is an adhesion molecule that plays an important role in inflammation. Protease inhibitors such as gabexate mesilate (GM) and ulinastatin (UTI) have been shown to improve the microcirculatory environment and reduce tissue damage, but the mechanism for this has yet to be fully elucidated. We investigated the effect of GM or UTI on EC regarding PAI-1 synthesis and ICAM-1 expression.


METHODS
Human umbilical vein endothelial cells (HUVECs) were obtained and stimulated with TNFalpha. GM or UTI was added to HUVECs just before TNFalpha stimulation. The PAI-1 activity in the culture medium of HUVECs was measured by using an enzymatic assay kit. The PAI-1 mRNA expression was assayed by a semiquantitative reverse transcriptase polymerase chain reaction assay. The ICAM-1 expression on HUVECs was assayed by a flow cytometric analysis.


RESULTS
GM inhibited the PAI-1 synthesis of HUVECs stimulated with TNFalpha in a dose-dependent manner as shown by the mRNA expression. However, UTI was not able to inhibit PAI-1 synthesis. In contrast, both GM and UTI significantly inhibited the ICAM-1 expression on HUVECs stimulated with TNFalpha.


CONCLUSIONS
These data suggest that GM may thus provide a beneficial effect which improves the microcirculatory environment and prevents tissue damage by inhibiting the activation of the vascular EC themselves.
Aosasa
S
S
Department of Surgery I, National Defense Medical College, Namiki 3-2, Tokorozawa Saitama, 359-8513, Japan.
Ono
S
S
Seki
S
S
Takayama
E
E
Tadakuma
T
T
Hiraide
H
H
Mochizuki
H
H
D001483:Base Sequence; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000502:physiology*; D016670:Gabexate / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006023:Glycoproteins / Q000494:pharmacology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism*; D008833:Microcirculation / Q000502:physiology; D017395:Plasminogen Activator Inhibitor 1 / Q000096:biosynthesis* / Q000235:genetics; D011480:Protease Inhibitors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
D017931:DNA Primers; D006023:Glycoproteins; D017395:Plasminogen Activator Inhibitor 1; D011480:Protease Inhibitors; D012333:RNA, Messenger; D011994:Recombinant Proteins; D014409:Tumor Necrosis Factor-alpha; D018799:Intercellular Adhesion Molecule-1; D016670:Gabexate; C028665:urinastatin