We investigated whether administration of estradiol to male mice augments mobilization of bone marrow-derived endothelial progenitor cells (EPC) and incorporation into foci of neovascularization after hind-limb ischemia, thereby contributing to blood flow restoration. Mice were randomized and implanted with placebo pellets or pellets containing low-dose estradiol (0.39 mg) or high-dose estradiol (1.7 mg). Hind-limb ischemia was induced by unilateral resection of the left femoral artery 1 week after pellet implantation, then EPC mobilization and functional recovery was evaluated. EPC recruitment was assessed in mice transplanted with bone marrow from transgenic donors expressing β-galactosidase driven by the Tie-2 promoter. EPC culture assay performed 2 weeks after pellet implantation revealed a significantly greater (p < 0.05) number of circulating EPCs in the high-dose estradiol group than in the low-dose estradiol and placebo groups. At 3 and 4 weeks after induction of hind-limb ischemia, perfusion was significantly greater (p < 0.05) in high-dose estradiol mice than in mice implanted with the low-dose estradiol or placebo pellets. At 1 and 4 weeks after hind-limb ischemia surgery, more bone marrow-derived EPCs, identified as β-galactosidase-positive cells, were observed in ischemic regions from high-dose estradiol animals than in low-dose (p < 0.05) or placebo groups (p < 0.05). These results indicate that estradiol dose-dependently increases the levels of EPCs in peripheral blood in male animals, improves the recovery of blood flow, and decreases limb necrosis after hind-limb ischemia, and that this enhancement occurs, in part, through augmentation of EPC mobilization and greater incorporation of bone marrow-derived EPCs into foci of neovascularization.
1 Asahara T Murohara T Sullivan A . Isolation of putative progenitor endothelial cells for angiogenesis. Science1997; 275: 964–967.
2.
2 Amrani DL Port S . Cardiovascular disease: potential impact of stem cell therapy. Expert Rev Cardiovasc Ther2003; 1: 453–461.
3.
3 Dobert N Britten M Assmus B . Transplantation of progenitor cells after reperfused acute myocardial infarction: evaluation of perfusion and myocardial viability with FDG-PET and thallium SPECT. Eur J Nucl Med Mol Imaging2004; 31: 1146–1151.
4.
4 Iwaguro H Yamaguchi J Kalka C . Endothelial progenitor cell vascular endothelial growth factor gene transfer for vascular regeneration. Circulation2002; 105: 732–738.
5.
5 Iwakura A Luedemann C Shastry S . Estrogen-mediated, endothelial nitric oxide synthase-dependent mobilization of bone marrow-derived endothelial progenitor cells contributes to reendothelialization after arterial injury. Circulation2003; 108: 3115–3121.
6.
6 Kudo FA Nishibe T Nishibe M Yasuda K . Autologous transplantation of peripheral blood endothelial progenitor cells (CD34+) for therapeutic angiogenesis in patients with critical limb ischemia. Int Angiol2003; 22: 344–348.
7.
7 Krasinski K Spyridopoulos I Asahara T van der Zee R Isner JM Losordo DW . Estradiol accelerates functional endothelial recovery after arterial injury. Circulation1997; 95: 1768–1772.
8.
8 Reyes M Dudek A Jahagirdar B Koodie L Marker PH Verfaillie CM . Origin of endothelial progenitors in human postnatal bone marrow. J Clin Invest2002; 109: 337–346.
9.
9 Shi Q Rafii S Wu MH . Evidence for circulating bone marrow-derived endothelial cells. Blood1998; 92: 362–367.
10.
10 Takahashi T Kalka C Masuda H . Ischemia- and cytokine-induced mobilization of bone marrow-derived endothelial progenitor cells for neovascularization. Nat Med1999; 5: 434–438.
11.
11 Hamada H Kim MK Iwakura A . Estrogen receptors alpha and beta mediate contribution of bone marrow-derived endothelial progenitor cells to functional recovery after myocardial infarction. Circulation2006; 114: 2261–2270.
12.
12 Iwakura A Shastry S Luedemann C . Estradiol enhances recovery after myocardial infarction by augmenting incorporation of bone marrow-derived endothelial progenitor cells into sites of ischemia-induced neovascularization via endothelial nitric oxide synthase-mediated activation of matrix metalloproteinase-9. Circulation2006; 113: 1605–1614.
13.
13 Shifren JL Tseng JF Zaloudek CJ . Ovarian steroid regulation of vascular endothelial growth factor in the human endometrium: implications for angiogenesis during the menstrual cycle and in the pathogenesis of endometriosis. J Clin Endocrinol Metab1996; 81: 3112–3118.
14.
14 Findlay JK . Angiogenesis in reproductive tissues. J Endocrinol1986; 111: 357–366.
15.
15 Koos RD Le Maire WJ . Factors that may regulate the growth and regression of blood vessels in the ovary. Semin Reprod Endocrinol1983; 1: 295–307.
16.
16 Reynolds LP Killielea SD Redmer DA . Angiogenesis in the female reproductive system. FASEB J1992; 6: 886–892.
17.
17 Goede V Schmidt T Kimmina S Kozian D Augustin HG . Analysis of blood vessel maturation process during cyclic ovarian angiogenesis. Lab Invest1998; 78: 1385–1394.
18.
18 Losordo DW Isner JM . Estrogen and angiogenesis: a review. Arterioscler Thromb Vasc Biol2001; 21: 6–12.
19.
19 Garcia PM Gimenez J Bonacasa B . 17beta-estradiol exerts a beneficial effect on coronary vascular remodeling in the early stages of hypertension in spontaneously hypertensive rats. Menopause2005; 12: 453–459.
20.
20 Miller AP Feng W Xing D . Estrogen modulates inflammatory mediator expression and neutrophil chemotaxis in injured arteries. Circulation2004; 110: 1664–1669.
21.
21 Patten RD Pourati I Aronovitz MJ . 17beta-estradiol reduces cardiomyocyte apoptosis in vivo and in vitro via activation of phospho-inositide-3 kinase/Akt signaling. Circ Res2004; 95: 692–699.
22.
22 Strehlow K Werner N Berweiler J . Estrogen increases bone marrow-derived endothelial progenitor cell production and diminishes neointima formation. Circulation2003; 107: 3059–3065.
23.
23 Goukassian DA Qin G Dolan C . Tumor necrosis factor-alpha receptor p75 is required in ischemia-induced neovascularization. Circulation2007; 115: 752–762.
24.
24 Asahara T Takahashi T Masuda H . VEGF contributes to postnatal neovascularization by mobilizing bone marrow-derived endothelial progenitor cells. EMBO J1999; 18: 3964–3972.
25.
25 Hirsch AT Criqui MH Treat-Jacobson D . Peripheral arterial disease detection, awareness, and treatment in primary care. JAMA2001; 286: 1317–1324.
26.
26 Criqui MH Vargas V Denenberg JO . Ethnicity and peripheral arterial disease: the San Diego Population Study. Circulation2005; 112: 2703–2707.
27.
27 Selvin E Erlinger TP . Prevalence of and risk factors for peripheral arterial disease in the United States: results from the National Health and Nutrition Examination Survey, 1999–2000. Circulation2004; 110: 738–743.
28.
28 Mendelsohn ME Karas RH . The protective effects of estrogen on the cardiovascular system. N Engl J Med1999; 340: 1801–1811.
29.
29 Nikolic I Liu D Bell JA Collins J Steenbergen C Murphy E . Treatment with an estrogen receptor-beta-selective agonist is cardioprotective. J Mol Cell Cardiol2007; 42: 769–780.
30.
30 Urata Y Ihara Y Murata H . 17Beta-estradiol protects against oxidative stress-induced cell death through the glutathione/glutaredoxin-dependent redox regulation of Akt in myocardiac H9c2 cells. J Biol Chem2006; 281: 13092–13102.
31.
31 Shinohara T Takahashi N Ooie T . Estrogen inhibits hyperthermia-induced expression of heat-shock protein 72 and cardioprotection against ischemia/reperfusion injury in female rat heart. J Mol Cell Cardiol2004; 37: 1053–1061.
32.
32 Zhai P Eurell TE Cotthaus R Jeffery EH Bahr JM Gross DR . Effect of estrogen on global myocardial ischemia-reperfusion injury in female rats. Am J Physiol2000; 279: H2766–H2775.
33.
33 Beer S Reincke M Kral M . High-dose 17beta-estradiol treatment prevents development of heart failure post-myocardial infarction in the rat. Basic Res Cardiol2007; 102: 9–18.
34.
34 Satoh M Matter CM Ogita H . Inhibition of apoptosis-regulated signaling kinase-1 and prevention of congestive heart failure by estrogen. Circulation2007; 115: 3197–3204.
35.
35 Kim JK Pedram A Razandi M Levin ER . Estrogen prevents cardiomyocyte apoptosis through inhibition of reactive oxygen species and differential regulation of p38 kinase isoforms. J Biol Chem2006; 281: 6760–6767.
36.
36 Hopkins SP Bulgrin JP Sims RL Bowman B Donovan DL Schmidt SP . Controlled delivery of vascular endothelial growth factor promotes neovascularization and maintains limb function in a rabbit model of ischemia. J Vasc Surg1998; 27: 886–894 discussion 95.
37.
37 Kyriakides ZS Petinakis P Kaklamanis L . Gender does not influence angiogenesis and arteriogenesis in a rabbit model of chronic hind limb ischemia. Int J Cardiol2003; 92: 83–91.
38.
38 Kyriakides ZS Petinakis P Kaklamanis L . Intramuscular administration of estrogen may promote angiogenesis and perfusion in a rabbit model of chronic limb ischemia. Cardiovasc Res2001; 49: 626–633.
39.
39 Senthilkumar A Smith RD Khitha J . Sildenafil promotes ischemia-induced angiogenesis through a PKG-dependent pathway. Arterioscler Thromb Vasc Biol2007; 27: 1947–1954.
40.
40 Tiidus PM Deller M Bombardier E Gul M Liu XL . Estrogen supplementation failed to attenuate biochemical indices of neutrophil infiltration or damage in rat skeletal muscles following ischemia. Biol Res2005; 38: 213–223.
41.
41 Di Stefano R Santoni T Barsotti MC . Different growth conditions for peripheral blood endothelial progenitors. Cardiovasc Radiat Med2002; 3: 172–175.
42.
42 Kalka C Masuda H Takahashi T . Vascular endothelial growth factor(165) gene transfer augments circulating endothelial progenitor cells in human subjects. Circ Res2000; 86: 1198–1202.
43.
43 Silvestre JS Tamarat R Ebrahimian TG . Vascular endothelial growth factor-B promotes in vivo angiogenesis. Circ Res2003; 93: 114–123.
44.
44 Yoon YS Losordo DW . All in the family: VEGF-B joins the ranks of proangiogenic cytokines. Circ Res2003; 93: 87–90.
45.
45 Yamaguchi J Kusano KF Masuo O . Stromal cell-derived factor-1 effects on ex vivo expanded endothelial progenitor cell recruitment for ischemic neovascularization. Circulation2003; 107: 1322–1328.
46.
46 Llevadot J Murasawa S Kureishi Y . HMG-CoA reductase inhibitor mobilizes bone marrow-derived endothelial progenitor cells. J Clin Invest2001; 108: 399–405.
47.
47 Bahlmann FH De Groot K Spandau JM . Erythropoietin regulates endothelial progenitor cells. Blood2004; 103: 921–926.
48.
48 Heeschen C Aicher A Lehmann R . Erythropoietin is a potent physiologic stimulus for endothelial progenitor cell mobilization. Blood2003; 102: 1340–1346.
49.
49 Westenbrink BD Lipsic E van der Meer P . Erythropoietin improves cardiac function through endothelial progenitor cell and vascular endothelial growth factor mediated neovascularization. Eur Heart J2007; 28: 2018–2027.
50.
50 Booth EA Obeid NR Lucchesi BR . Activation of estrogen receptor-alpha protects the in vivo rabbit heart from ischemia-reperfusion injury. Am J Physiol2005; 289: H2039–H2047.
