The inhibition of tube formation in a collagen–fibrinogen, three-dimensional gel by cleaved kininogen (HKa) and HK domain 5 (D5) is dependent on Src family kinases

Liu, Yuchuan; Sáinz, Irma M.; Yu, Wu; Pixley, Robin A.; Espinola, Ricardo; Hassan, Sarmina; Khan, Mohammad M.; Colman, Robert W.

doi:10.1016/j.yexcr.2007.10.008

Cited by 20 publications

(34 citation statements)

References 56 publications

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“…On reduced gels, HKa possesses a heavy chain (M r ϭ 62 kDa) and a light chain (Mr ϭ 45 kDa). Endotoxin in HKa was measured and removed as described in a previous publication (40). Only HKa was used in these experiments; no intact high-molecular kininogen (HK) or domain 5 (D5) was used in this study.…”

Section: Methodsmentioning

confidence: 99%

The inhibitory effect of HKa in endothelial cell tube formation is mediated by disrupting the uPA-uPAR complex and inhibiting its signaling and internalization

Liu¹,

Cao²,

Sáinz³

et al. 2008

American Journal of Physiology-Cell Physiology

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In two-dimensional (2-D) culture systems, we have previously shown that cleaved two-chain high-molecular-weight kininogen (HKa) or its domain 5 induced apoptosis by disrupting urokinase plasminogen activator (uPA) receptor (uPAR)-integrin signal complex formation. In the present study, we used a three-dimensional (3-D) collagen-fibrinogen culture system to monitor the effects of HKa on tube formation. In a 3-D system, HKa significantly inhibited tube and vacuole formation as low as 10 nM, which represents 1.5% of the physiological concentration of high-molecular-weigh kininogen (660 nM), without apparent apoptosis. However, HKa (300 nM) completely inhibited tube formation and increased apoptotic cells about 2-fold by 20-24 h of incubation. uPA-dependent ERK activation and uPAR internalization regulate cell survival and migration. In a 2-D system, we found that exogenous uPA-induced ERK phosphorylation and uPAR internalization were blocked by HKa. In a 3-D system, we found that not only uPA-uPAR association but also the activation of ERK were inhibited by HKa. HKa disrupts the uPA-uPAR complex, inhibiting the signaling pathways, and also inhibits uPAR internalization and regeneration to the cell surface, thereby interfering with uPAR-mediated cell migration, proliferation, and survival. Thus, our data suggest that the suppression of ERK activation and uPAR internalization by HKa contributes to the inhibition of tube formation. We conclude that in this 3-D collagen-fibrinogen gel, HKa modulates the multiple functions of uPAR in endothelial cell tube formation, a process that is closely related to in vivo angiogenesis.

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Section: Methodsmentioning

confidence: 99%

The inhibitory effect of HKa in endothelial cell tube formation is mediated by disrupting the uPA-uPAR complex and inhibiting its signaling and internalization

Liu¹,

Cao²,

Sáinz³

et al. 2008

American Journal of Physiology-Cell Physiology

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“…The ability of endothelial cells to assemble into vessel-like structures (tubes) is an important aspect of blood vessel formation that is sensitive to inhibition by HKa (23). To investigate whether ferritin blocks the inhibitory effect of HKa on tube formation, endothelial cells were incubated with matrigel, which induces the cells to assemble into branching tube-like structures (24,25), in the presence of HKa, ferritin, or the combination of HKa plus ferritin.…”

Section: Ferritin Restores the Ability Of Hka-treated Endothelial Celmentioning

confidence: 99%

Regulatory effects of ferritin on angiogenesis

Coffman

Parsonage²,

D’Agostino

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

138

118

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Angiogenesis, the synthesis of new blood vessels from preexisting vessels, plays a critical role in normal wound healing and tumor growth. HKa (cleaved high molecular weight kininogen) is an endogenous inhibitor of angiogenesis formed by the cleavage of kininogen on endothelial cells. Ferritin is a protein principally known for its central role in iron storage. Here, we demonstrate that ferritin binds to HKa with high affinity (K d 13 nM). Further, ferritin antagonizes the antiangiogenic effects of HKa, enhancing the migration, assembly, and survival of HKa-treated endothelial cells. Effects of ferritin were independent of its iron content. Peptide mapping revealed that ferritin binds to a 22-aa subdomain of HKa that is critical to its antiangiogenic activity. In vivo, ferritin opposed HKa's antiangiogenic effects in a human prostate cancer xenograft, restoring tumordependent vessel growth. Ferritin-mediated regulation of angiogenesis represents a new angiogenic regulatory pathway, and identifies a new role for ferritin in cell biology.iron ͉ kininogen

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“…[16] Csk inhibits Lck activity, therefore inhibition of Csk leads to a potentiation of Lck activity. [22,23] The use of PP2 as a tool to selectively address Lck activity next to Csk activity indicates that our observations might be fully attributed to the action of PP2 on Lck. [23] Nevertheless, it cannot be excluded that the balance of the activities of these two kinases or an unknown secondary target and therefore also the sensitivity to inhibition is a function of the nature of the stimulus.…”

Section: Discussionmentioning

confidence: 86%

Stimulus Dependence of the Action of Small‐Molecule Inhibitors in the CD3/CD28 Signalling Network

et al. 2008

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Cells in the body are exposed simultaneously to a multitude of various signals. Inside a cell, molecular signalling networks integrate this information into a physiologically meaningful response. Interestingly, in the cellular testing of drug candidates, this complexity is largely ignored. Compounds are tested for cells that are challenged with one stimulus only. The activation of T lymphocytes through engagement of the T cell receptor (TCR)-CD3 complex and CD28 coreceptor is a prominent example for a cellular response that depends on the integration of signals. We investigated the cellular response characteristics of this network at different strengths of receptor and coreceptor activation. A novel cellular microarray-based approach, in which various combinations of antibodies directed against the CD3 complex and CD28 were spotted, was employed for analysing the stimulus dependence of activation of the transcription factor NFAT and actin reorganisation. For both responses, quantitative differences in inhibitor activity were observed. Remarkably, for IL-2 expression, which was detected by standard ELISA, low doses of the Src-family kinase inhibitor PP2 strongly potentiated IL-2 expression at high-level, but not at low-level, CD28 co-engagement. Therefore, for a physiologically highly relevant signalling network, the cellular response might vary qualitatively with only quantitative variations of a stimulus. This level of complexity should be considered in early cellular drug testing.

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The inhibition of tube formation in a collagen–fibrinogen, three-dimensional gel by cleaved kininogen (HKa) and HK domain 5 (D5) is dependent on Src family kinases

Cited by 20 publications

References 56 publications

The inhibitory effect of HKa in endothelial cell tube formation is mediated by disrupting the uPA-uPAR complex and inhibiting its signaling and internalization

The inhibitory effect of HKa in endothelial cell tube formation is mediated by disrupting the uPA-uPAR complex and inhibiting its signaling and internalization

Regulatory effects of ferritin on angiogenesis

Stimulus Dependence of the Action of Small‐Molecule Inhibitors in the CD3/CD28 Signalling Network

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