Critical function of Bmx/Etk in ischemia-mediated arteriogenesis and angiogenesis
Yun He, … , Kari Alitalo, Wang Min
Yun He, … , Kari Alitalo, Wang Min
Published September 1, 2006
Citation Information: J Clin Invest. 2006;116(9):2344-2355. https://doi.org/10.1172/JCI28123.
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Research Article Angiogenesis

Critical function of Bmx/Etk in ischemia-mediated arteriogenesis and angiogenesis

Abstract

Bmx/Etk non-receptor tyrosine protein kinase has been implicated in endothelial cell migration and tube formation in vitro. However, the role of Bmx in vivo is not known. Bmx is highly induced in the vasculature of ischemic hind limbs. We used both mice with a genetic deletion of Bmx (Bmx-KO mice) and transgenic mice expressing a constitutively active form of Bmx under the endothelial Tie-2 enhancer/promoter (Bmx-SK-Tg mice) to study the role of Bmx in ischemia-mediated arteriogenesis/angiogenesis. In response to ischemia, Bmx-KO mice had markedly reduced, whereas Bmx-SK-Tg mice had enhanced, clinical recovery, limb perfusion, and ischemic reserve capacity when compared with nontransgenic control mice. The functional outcomes in these mice were correlated with ischemia-initiated arteriogenesis, capillary formation, and vessel maturation as well as Bmx-dependent expression/activation of TNF receptor 2– and VEGFR2-mediated (TNFR2/VEGFR2-mediated) angiogenic signaling in both hind limb and bone marrow. More importantly, results of bone marrow transplantation studies showed that Bmx in bone marrow–derived cells plays a critical role in the early phase of ischemic tissue remodeling. Our study provides the first demonstration to our knowledge that Bmx in endothelium and bone marrow plays a critical role in arteriogenesis/angiogenesis in vivo and suggests that Bmx may be a novel target for the treatment of vascular diseases such as coronary artery disease and peripheral arterial disease.

Authors

Yun He, Yan Luo, Shibo Tang, Iiro Rajantie, Petri Salven, Matthias Heil, Rong Zhang, Dianhong Luo, Xianghong Li, Hongbo Chi, Jun Yu, Peter Carmeliet, Wolfgang Schaper, Albert J. Sinusas, William C. Sessa, Kari Alitalo, Wang Min

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Figure 2

EC-specific transgenic mice: specific expression of Bmx-SK in ECs driven byTie-2 promoter.

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EC-specific transgenic mice: specific expression of B...
(A) Schema of transgenic vector showing inserted Flag-tagged human Bmx-SK and location of primers used for genotyping by PCR. (B) Genotyping for Bmx-SK transgene. Tail genomic DNA was isolated, and genotyping was performed by PCR with specific primers as described in Methods. Lines 4 and line 6 were positive for Bmx-SK. (C) Tissue distribution of Bmx-SK expression. Bmx-SK mRNA in indicated tissues of Bmx-SK-Tg mice (n = 2 for each line) was quantified by qRT-PCR with 18S rRNA for normalization. Data are presented fold increase, with expression of Bmx-SK mRNA in liver taken as 1.0. Ar, aorta; Br, brain; H, heart; Li, liver; Lu, lung; Mu, muscle. (D) Bmx-SK expression in muscle of Bmx-SK-Tg mice (Tg lines 4 and 6). Mouse lower hind limb was collected and homogenized. Bmx-SK was detected by Western blotting with anti-Flag antibody. C57BL/6 muscle tissue (WT) was used as a negative control. β-Tubulin was used for protein normalization. (E) Relative expression level of Bmx-SK and endogenous Bmx. Expression of both transgenic and endogenous Bmx in hind limb was determined by Western blotting with anti-Bmx. Bmx, Bmx-SK, and nonspecific bands (ns) are indicated. (F) Bmx-SK expression in muscle capillaries of Bmx-SK-Tg mice. Mouse lower hind limb was collected as frozen sections, and Bmx-SK and EC markers were detected by immunohistochemistry with anti-Bmx and anti-CD31 antibodies, respectively. Bmx-SK–positive capillaries are indicated by arrows. (G) C57BL/6 (WT) and Bmx-KO (KO) muscle tissues were used as negative and a positive controls, respectively.