Background-Many angiogenic factors require endothelium-derived nitric oxide (NO) to exert their effects. Recently, an endogenous competitive antagonist of NO synthase has been characterized: asymmetric dimethylarginine (ADMA). Elevated plasma levels of ADMA reduce NO synthesis in hypercholesterolemia. Accordingly, we hypothesized that hypercholesterolemia impairs angiogenesis by an ADMA-dependent mechanism. Methods and Results-Angiogenesis was assessed with the use of a disk angiogenesis system implanted subcutaneously in normal (E ϩ ) mice or apolipoprotein (apo)E-deficient hypercholesterolemic (E Ϫ ) mice. After 2 weeks, the disks were removed, and the fibrovascular growth area was used as an index of angiogenesis. Basal and fibroblast growth factor-stimulated angiogenesis was impaired in E Ϫ mice, associated with an elevation in plasma ADMA. Oral administration of L-arginine reversed the impairment of angiogenesis in E Ϫ mice. By contrast, oral administration of

Background-Developmentally regulated endothelial locus-1 (Del-1) is an extracellular matrix protein that is expressed by endothelial cells during embryological vascular development. We speculated that Del-1 may be reexpressed in ischemia and may be involved in endogenous angiogenesis. Methods and Results-Del-1 protein was detected by immunohistochemistry in murine ischemic hindlimb after femoral artery excision. To determine whether exogenous Del-1 would augment angiogenesis in vivo, Del-1 or vehicle was administered for 3 weeks by intramuscular injection of murine ischemic hindlimbs. Angiogenesis was quantified by gadolinium-MRI perfusion and capillary densitometry. We used a disc angiogenesis system (DAS) to characterize the angiogenic response to vehicle (PBS), Del-1, Del-1 mutant (altered RGD domain), Del-1 minor (truncated discoidin-I-like domain), or basic fibroblast growth factor. After 14 days, the discs were extracted and sectioned to quantify vascular growth by morphometry. Endogenous Del-1 protein expression was increased in ischemic hindlimbs. Administration of Del-1 increased hindlimb vascular flow index and capillary density. In the DAS, Del-1 doubled fibrovascular growth, as did basic fibroblast growth factor. However, angiogenesis was not enhanced by the Del-1 mutant or Del-1 minor proteins. Conclusions-Del-1 is expressed in ischemic tissue. Del-1 stimulates angiogenesis, an effect that is dependent on the RGD motif and a second signaling sequence in the discoidin-I-like domain. Exogenous intramuscular administration of Del-1 significantly enhances angiogenesis in the murine ischemic hindlimb. Del-1 may prove to be a novel therapeutic agent for patients with ischemia.

We tested whether supplementation with L-arginine can augment aerobic capacity, particularly in conditions where endothelium-derived nitric oxide (EDNO) activity is reduced. Eight-week-old wild-type (E(+)) and apolipoprotein E-deficient mice (E(-)) were divided into six groups; two groups (LE(+) and LE(-)) were given L-arginine (6% in drinking water), two were given D-arginine (DE(+) and DE(-)), and two control groups (NE(+) and NE(-)) received no arginine supplementation. At 12-16 wk of age, the mice were treadmill tested, and urine was collected after exercise for determination of EDNO production. NE(-) mice demonstrated a reduced aerobic capacity compared with NE(+) controls [maximal oxygen uptake (VO(2 max)) of NE(-) = 110 +/- 2 (SE) vs. NE(+) = 122 +/- 3 ml O(2). min(-1). kg(-1), P < 0.001]. This decline in aerobic capacity was associated with a diminished postexercise urinary nitrate excretion. Mice given L-arginine demonstrated an increase in postexercise urinary nitrate excretion and aerobic capacity in both groups (VO(2 max) of LE(-) = 120 +/- 1 ml O(2). min(-1). kg(-1), P < 0.05 vs. NE(-); VO(2 max) of LE(+) = 133 +/- 4 ml O(2). min(-1). kg(-1), P < 0.01 vs. NE(+)). Mice administered D-arginine demonstrated an intermediate increase in aerobic capacity in both groups. We conclude that administration of L-arginine restores exercise-induced EDNO synthesis and normalizes aerobic capacity in hypercholesterolemic mice. In normal mice, L-arginine enhances exercise-induced EDNO synthesis and aerobic capacity.