Tissue kallikrein is synthesized and secreted by human vascular endothelial cells

Abstract

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.

Angiogenesis in cervical cancer is mediated by HeLa metabolites through endothelial cell tissue kallikrein

“…This increase appeared to be in direct response to tumour metabolite concentration in the conditioned-medium, and suggests functional activation of pre-kallikrein. This latter effect is supported by the studies of Shih and co-workers who showed that ovarian cancers secrete kallikreins 6, 7, 8 and 10 (47) and the Yayama group who have shown that activated TK is continually secreted by HUVECS (20). Further, in the present study, HeLa metabolites caused significant increases in HUVEC proliferation, by as much as 200%.…”

“…The involvement of the KKS in angiogenesis has been demonstrated by the expression of TK in bovine angiogenic endothelial cells (19,20), non-angiogenic HUVECs and human coronary artery endothelial cells (20). In this study, we also demonstrate the presence of TK and TK mRNA in angiogenic HUVECs, but this is the first demonstration of the ability of metabolites from cervical cancer cells to induce the release of TK from HUVECs.…”

“…Our anti-TK IgG detected both the prekallikrein stored within unchallenged HUVECs as well as TK released from HUVECs following activation by cervical cancer cell metabolites. Although other studies, using PCR, amidolytic activity and receptor antagonists have demonstrated the release of active TK by human vascular endothelial cells (20), none have demonstrated a dosedependent release due to tumour metabolites. Here, we have been able to show, by ELISA, a significant increase in total kallikrein content in the spent medium of challenged HUVECs.…”

“…Some pro-angiogenic factors shown to be involved in cervical cancer include vascular endothelial growth factor (VEGF) (14)(15)(16) and platelet-derived growth factor (PDGF) (17). However, there are no studies on the role of tissue kallikrein (TK) in cervical cancer angiogenesis even though TK and its gene, KLK1 (EC 3.4.21.35) have been shown to induce angiogenesis in an ischaemic hind-limb model (18) and to be present in the endothelial cells of human umbilical cord vein (HUVEC) (19) and human coronary artery (20).…”

“…Other functions of TK include the proteolytic breakdown of progelatinase and pro-collagenase (21) with concomitant destabilisation of the stroma, thereby promoting the migration of both endothelial and tumour cells (22). Although TK and KLK1 have been demonstrated in endothelial cells (19,20) and in tumours of the human stomach (23), oesophagus (24) and breast (25), there are no studies on the role of the kallikrein-kinin system (KKS) in the pathogenesis of cervical cancer. Components of the KKS are also considered to form the first line onset of inflammation (26), a process described as important in the pathogenesis of cervical cancer (27,28), and the vascularity observed in cervical cancer (29).…”