doi: 10.1161/ATVBAHA.107.158949
© 2008 American Heart Association, Inc.
Letters to the Editor |
The Puzzling Role of TRAIL in Endothelial Cell Biology
Division of Biotechnology Discovery Research, Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Ind
In Response:
The letter by Secchiero and Zauli regarding our study on TRAIL suppression via the Egr-1/Erk1/2 pathway by activated protein C (APC)1 comments positively on the mechanistic data, but questions the pathophysiological role of TRAIL on endothelial cells, largely based on their inability to detect TRAIL in cultured human umbilical vein endothelial cells (HUVECs). In our article, we used HUVECs as a model system to study the signaling response to APC, with TRAIL as a response gene after activation by tumor necrosis factor (TNF)-alpha. The expression of TRAIL by HUVECs and its regulation by inflammatory mediators is well described in the literature. As we originally cited, Fu et al2 demonstrated that TRAIL is highly expressed in cultured HUVECs and regulated by overexpression of Egr-1. Viemann and colleagues3 demonstrated TRAIL expression in HUVECs, which could be upregulated 4-fold by TNF-
. Larghero et al4 studied the effect of lipoic acid on HUVEC function and demonstrated an induction of TRAIL expression and suggested a relationship to in vivo antiangiogenic activity. Warke et al5 used both ELISA and flow cytometry to demonstrate that HUVECs can be induced to express TRAIL. In other cultured endothelial cells, TRAIL expression also has been documented; eg, Prat et al6 demonstrated that human brain endothelial cells express TRAIL, which is dramatically upregulated in response to cytokines, and Pritzker et al7 were able to immunoprecipitate TRAIL protein from human microvascular endothelial cells. As additional support for the physiological relevance of TRAIL expression beyond cultured cells, Spierings et al8 demonstrated TRAIL expression in human lung vascular endothelial cells by immunohistochemistry. Similarly, Favre and colleagues,9 using an approach to directly isolate lung capillary endothelial cells at high purity, demonstrated that TRAIL was not only expressed but was expressed at a relatively high ratio compared with unfractionated lung. Finally, using immunohistochemical analysis, expression of TRAIL protein has been shown in the endothelium of the human cornea.10 Overall, the data we reported are consistent with these published reports in HUVECs and other cultured endothelial cells showing both TRAIL expression and regulation by inflammatory mediators. In light of these articles showing TRAIL expression in endothelial cells, we do not have an explanation for the lack of expression by flow analysis described in the letter by Secchiero and Zauli. There are myriad reasons for a negative study, and in this case some possible explanations could be different source/preparation of the HUVECs, different culture conditions, or loss of surface TRAIL in preparing monolayers for flow analysis.
Secchiero and Zauli also comment that we postulated a detrimental role for TRAIL in vascular biology, citing studies showing that particular culture conditions are required to show TRAIL-mediated cytotoxicity, whereas other conditions in cell culture show a protective effect of TRAIL. In addition to the association between modulation of inflammatory mediators and endogenous TRAIL induction shown in O’Brien et al,1 we see a significant effect of exogenous TRAIL on the activation of endothelial cell caspase-3 using standard HUVEC growth conditions (Figure). Of interest, Kirsch et al11 recently demonstrated that human microvascular endothelial cells express TRAIL, which is negatively regulated by coincubation with apoptotic or necrotic endothelial cells. Further, Chen and Easton12 showed that TRAIL increases neutrophil adhesion when it concurrently induces apoptosis both in EA.hy926 endothelial cells and in sensitized HUVECs. Additional recent studies4,13 also have suggested that TRAIL can have apoptotic functions as well as antiangiogenic and inflammatory functions, consistent with the data we reported.
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It was not our intention to suggest, nor did we make a generalization, that TRAIL has a detrimental effect in vascular biology. As with most regulatory molecules, a biological effect being detrimental versus beneficial must be made in the context of the physiology or pathophysiology being examined. Clearly, in some instances, vascular apoptosis and antiangiogenesis can be detrimental (revascularization), whereas in other instances it could have a beneficial effect (tumor inhibition). In endothelial dysfunction as occurs in severe sepsis, increased inflammatory activity and apoptosis of the endothelium is undesirable.
In summary, the broader intent of the article was to describe a novel signaling pathway for APC, using the well documented regulation of TRAIL expression in HUVECs as a model system. Regarding the comment on the potential usefulness of a drug modulating endothelial TRAIL, as we stated in the article, the relative role of the EPCR-independent signaling path will require further study. We agree with Secchiero and Zauli that continued efforts are warranted to dissect the potential role of TRAIL in different physiological contexts.
Acknowledgments
Disclosures
The authors are employees of Eli Lilly and Co, who produces human APC for treatment of severe sepsis.
References
1. O’Brien LA, Richardson MA, Mehrbod S, Berg DT, Gerlitz B, Gupta A, Grinnell BW. Activated protein C decreases tumor necrosis factor related apoptosis-inducing ligand by an EPCR independent mechanism involving Egr-1/Erk-1/2 activation. Arterioscler Thromb Vasc Biol. 2007; 27: 2634–41.
2. Fu M, Zhu X, Zhang J, Liang J, Lin Y, Zhao L, Ehrengruber MU, Chen YE. Egr-1 target genes in human endothelial cells identified by microarray analysis. Gene. 2003; 315: 33–41:34:27.[CrossRef][Medline] [Order article via Infotrieve]
3. Viemann D, Goebeler M, Schmid S, Nordhues U, Klimmek K, Sorg C, Roth J. TNF induces distinct gene expression programs in microvascular and macrovascular human endothelial cells. J Leukoc Biol. 2006; 80: 174–185.
4. Larghero P, Venè R, Minghelli S, Travaini G, Morini M, Ferrari N, Pfeffer U, Noonan D, Albini A, Benelli R. Biological assays and genomic analysis reveal lipoic acid modulation of endothelial cell behavior and gene expression. Carcinogenesis. 2007; 28: 1008–1020.
5. Warke RV, Martin KJ, Giaya K, Shaw SK, Rothman AL, Bosch I. TRAIL is a novel anti-viral protein against dengue virus. J Virol. In press.
6. Prat A, Biernacki K, Wosik K, Antel J. Glial cell influence on the human blood-brain barrier. Glia. 2001; 36: 145–155.[CrossRef][Medline] [Order article via Infotrieve]
7. Pritzker LB, Scatena M, Giachelli CM. The role of osteoprotegerin and tumor necrosis factor-related apoptosis-inducing ligand in human microvascular endothelial cell survival. Mol Biol Cell. 2004; 15: 2834–2841.
8. Spierings D, de Vries E, Vellenga E, van den Heuvel F, Koornstra J, Wesseling J, Hollema H, de Jong S. Tissue distribution of the death ligand TRAIL and its receptors. J Histochem Cytochem. 2004; 52: 821–831.
9. Favre C, Mancuso M, Maas K, McLean J, Baluk P, McDonald D. Expression of genes involved in vascular development and angiogenesis in endothelial cells of adult lung. Am J Physiol Heart Circ Physiol. 2003; 285: H1917–H1938.
10. Lee H, Herndon J, Barreiro R, Griffith T, Ferguson T. TRAIL: a mechanism of tumor surveillance in an immune privileged site. J Immunol. 2002; 169: 4739–4744.
11. Kirsch T, Woywodt A, Beese M, Wyss K, Park J, Erdbruegger U, Hertel B, Haller H, Haubitz M. Engulfment of apoptotic cells by microvascular endothelial cells induces proinflammatory responses. Blood. 2007; 109: 2854–2862.
12. Chen P, Easton A. Apoptotic phenotype alters the capacity of tumor necrosis factor-related apoptosis-inducing ligand to induce human vascular endothelial activation. J Vasc Res. 2007; 45: 111–122.[Medline] [Order article via Infotrieve]
13. Indraccolo S, Pfeffer U, Minuzzo S, Esposito G, Roni V, Mandruzzato S, Ferrari N, Anfosso L, Dell’Eva R, Noonan DM, Chieco-Bianchi L, Albini A, Amadori A. Identification of genes selectively regulated by IFNs in endothelial cells. J Immunol. 2007; Jan 15 178 (2): 1122–1135.
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