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Cathepsin L is required for endothelial progenitor cell–induced neovascularization

Abstract

Infusion of endothelial progenitor cells (EPC), but not of mature endothelial cells, promotes neovascularization after ischemia. We performed gene expression profiling of EPC and endothelial cells to identify genes that might be important for the neovascularization capacity of EPC. Notably, the protease cathepsin L (CathL) was highly expressed in EPC as opposed to endothelial cells and was essential for matrix degradation and invasion by EPC in vitro. CathL-deficient mice showed impaired functional recovery following hind limb ischemia, supporting the concept of a crucial role for CathL in postnatal neovascularization. Infused CathL-deficient progenitor cells neither homed to sites of ischemia nor augmented neovascularization. Forced expression of CathL in mature endothelial cells considerably enhanced their invasive activity and sufficed to confer their capacity for neovascularization in vivo. We concluded that CathL has a critical role in the integration of circulating EPC into ischemic tissue and is required for EPC-mediated neovascularization.

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Figure 1: Gene expression analysis of EPC and HUVEC.
Figure 2: The expression and activity of cathepsin L is increased in EPC.
Figure 3: Role of CathL in neovascularization.
Figure 4: Role of CathL in EPC invasion.
Figure 5: CathL is required for improvement of neovascularization.
Figure 6: Overexpression of CathL in mature endothelial cells partially rescued the impaired improvement of neovascularization.

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Acknowledgements

We are thankful to J. Gille (University of Frankfurt, Frankfurt, Germany) and Z. Werb (University of California, San Francisco, California) for providing the MMP9-deficient mice. DCG-04 was donated by M. Bogyo (Stanford University Medical School, California). We would like to thank A. Knau, M. Näher, and M. Muhly-Reinholz for technical help. This study was supported by the Deutsche Forschungsgemeinschaft (Di 600/4-1, He 3044/2-2 and VA151/4-3), the Alfried Krupp Stiftung (S.D.), the Fonds der Chemischen Industrie (T.R. and C.P.), by the Deutsche Krebshilfe (T.R.), and in part by National Institutes of Health Grant HL071954A (DE-AC03-76SF00098 (L.A.P.)). K.S was in part supported by the Japan Heart Foundation/Bayer Yakuhin Research Grant Abroad. The authors C.U., C.H., A.A., T.B., A.M.Z. and S.D. belong to the European Vascular Genomics Network (http://www.evgn.org) a Network of Excellence supported by the European Community's sixth Framework Programme for Research Priority 1 “Life sciences, genomics and biotechnology for health” (Contract number LSHM-CT-2003-503254).

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Correspondence to Stefanie Dimmeler.

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Supplementary information

Supplementary Fig. 1

Effect of lymphocyte infusion on neovascularization. (PDF 126 kb)

Supplementary Fig. 2

Expression of cathepsin L on different cell types. (PDF 36 kb)

Supplementary Fig. 3

Effect of cathepsin L on progenitor cell function. (PDF 23 kb)

Supplementary Fig. 4

Role of cathepsin L in mature endothelial cell function. (PDF 45 kb)

Supplementary Fig. 5

Proteolytic activity of EPC and progenitor cells in lysates and supernatants. (PDF 25 kb)

Supplementary Fig. 6

Confocal proteolysis assay. (PDF 73 kb)

Supplementary Fig. 7

Histological evaluation of tissue vascularization—capillary density. (PDF 317 kb)

Supplementary Fig. 8

Histological evaluation of tissue vascularization—conductance vessels. (PDF 59 kb)

Supplementary Fig. 9

Role of cathepsin L in progenitor cell–induced perfusion. (PDF 296 kb)

Supplementary Fig. 10

Incorporation of endothelial progenitor cells. (PDF 101 kb)

Supplementary Fig. 11

Role of cathepsin L in tumor vascularization. (PDF 417 kb)

Supplementary Fig. 12

Role of cathepsin L in incorporation of bone marrow cells. (PDF 63 kb)

Supplementary Fig. 13

MMP-9 and cathepsin L activity in ischemic tissues. (PDF 24 kb)

Supplementary Table 1

The mRNA expression (normalized data) of various proteases and protease inhibitors in EPC, HUVEC and monocytes is summarized. n = 3; *P < 0.05 versus HUVEC. (PDF 19 kb)

Supplementary Movie 1

Incorporation of endothelial progenitor cells (AVI 15480 kb)

Supplementary Methods (PDF 49 kb)

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Urbich, C., Heeschen, C., Aicher, A. et al. Cathepsin L is required for endothelial progenitor cell–induced neovascularization. Nat Med 11, 206–213 (2005). https://doi.org/10.1038/nm1182

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