Tissue kallikrein is synthesized and secreted by human vascular endothelial cells.
The generation of kinins on the surface of vascular endothelium has been postulated in two pathways involving plasma kallikrein and tissue kallikrein; the former pathway has been well documented, but the latter is controversial. To clarify the presence of a kinin-generating system on endothelium, we examined whether human umbilical vein endothelial cells (HUVEC) synthesize and release tissue kallikrein in vitro. Kallikrein-like activity hydrolyzing a peptide Pro-Phe-Arg-4-methyl-coumaryl-7-amide was detected in the culture medium of HUVEC and was inhibited by aprotinin but not by soybean trypsin inhibitor. Western blotting of HUVEC medium using anti-human tissue kallikrein antibodies demonstrated the release of tissue kallikrein from HUVEC, and the reverse transcription-polymerase chain reaction (RT-PCR) followed by Southern blotting revealed the expression of tissue kallikrein mRNA in HUVEC. HUVEC metabolically labeled with [35S]methionine released radioactive proteins corresponding to tissue kallikrein. RT-PCR also showed the expression of low-molecular-weight kininogen (L-kininogen) mRNA in HUVEC. The cGMP levels in HUVEC were significantly elevated by the incubation with angiotensin converting enzyme inhibitor, lisinopril, and the elevation was completely inhibited by aprotinin or bradykinin B2-receptor antagonist, FR172357. These results suggest that the endothelial cells continuously release an active form of tissue kallikrein which enables generation of kinins on the vascular endothelium.
Yayama
Katsutoshi
K
Department of Pharmacology, Faculty of Pharmaceutical Sciences and High Technology Research Center, Kobe Gakuin University, Ikawadani-cho, Nishi-ku, 651-2180, Kobe, Japan.
Kunimatsu
Naomi
N
Teranishi
Yumiko
Y
Takano
Masaoki
M
Okamoto
Hiroshi
H
D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D015153:Blotting, Western; D002478:Cells, Cultured; D002845:Chromatography; D017077:Culture Media, Conditioned; D006152:Cyclic GMP / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D007704:Kininogens / Q000032:analysis; D017706:Lisinopril / Q000494:pharmacology; D012333:RNA, Messenger / Q000032:analysis; D020840:Tissue Kallikreins / Q000032:analysis / Q000096:biosynthesis* / Q000737:chemistry
D000806:Angiotensin-Converting Enzyme Inhibitors; D017077:Culture Media, Conditioned; D007704:Kininogens; D012333:RNA, Messenger; D017706:Lisinopril; D020840:Tissue Kallikreins; D006152:Cyclic GMP