TRAINING INCREASES COLLATERAL BLOOD FLOW IN AGED RATS WITH FEMORAL ARTERY LIGATION

We tested the hypothesis that aged animals are as responsive as the young adult animals in expanding collateral vasculature under a similar treatment of basic fibroblast growth factor (bFGF). Two age groups of male Fischer 344 rats (11 mo old; n = 32, 23 mo old; n = 43) weighing ∼385 g were subdivided into normal, acute ligation [femoral artery (FA) ligated 3 days before blood flow (BF) measurement] or ligated groups for 16 days and received recombinant human bFGF intra-arterial infusion at doses of 0, 0.5, 5, and 50 μg ⋅ kg− 1 ⋅ day− 1. BF was determined with 85Sr- and 141Ce-labeled microspheres during treadmill running at 15 and 20 m/min at 15% grade. Blood presure (BP) values were ∼149 and ∼163 mmHg ( p < 0.05); heart rates were ∼496 and ∼512 beats/min in the aged and young adult groups during running, respectively. Maximal collateral BF values were confirmed by no additional BF increase in the calf muscle at the higher speed. Ligation of the FA for 3 days reduced the BF reserve to the calf muscle by ∼90%. Calf muscle BF was modestly greater (10 ml ⋅ min− 1 ⋅ 100 g− 1) by 16 days in the carrier group. bFGF infusion expanded collateral BF in a dose-dependent manner with an increase of 33 and 42 ml ⋅ min− 1 ⋅ 100 g− 1 ( P < 0.001) in the 5 and 50 μg ⋅ kg− 1 ⋅ day− 1bFGF groups, respectively. Aged animals showed similar BF improvements as observed with the adult groups in response to ligation surgery and bFGF treatment. Our data indicate that the aged rats (∼23 mo old) remain responsive to exogenous bFGF induced in developing collateral-dependent BF as the young adult (∼11 mo old) controls. This suggests that the influence of bFGF in expanding collateral BF should not be preempted in the aged group, the population most affected by peripheral arterial insufficiency.

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Abstract 15312: Genetic Deletion of Toll-like Receptor 9 Accelerates Blood Flow Recovery after Hindlimb Ischemia

Circulation ◽

10.1161/circ.130.suppl_2.15312 ◽

2014 ◽

Vol 130 (suppl_2) ◽

Author(s):

Sachiko Nishimoto ◽

Daiju Fukuda ◽

Yasutomi Higashikuni ◽

Kimie Tanaka ◽

Yoichiro Hirata ◽

...

Keyword(s):

Endothelial Cells ◽

Blood Flow ◽

Femoral Artery ◽

Cpg Odn ◽

Toll Like Receptor ◽

Artery Ligation ◽

Ischemic Limb ◽

Tnf Α ◽

Genetic Deletion ◽

Inflammatory Molecules

Background: Peripheral artery disease causes significant functional disability and results in impaired quality of life. Toll-like receptor (TLR)-2, 3 and 4 are suggested to participate in blood flow recovery in ischemic limb by modulating inflammation and angiogenesis, however, the role of TLR9 remains unknown. TLR9 recognizes bacterial unmethylated DNA and plays a role in innate defense, although it can also provoke inflammation in response to fragmented DNA released from regenerated mammalian cells. This study tested the hypothesis that genetic deletion of TLR9 accelerates blood flow recovery after femoral artery ligation by inhibiting inflammation and improving endothelial cell function. Methods and Results: Unilateral femoral artery ligation was performed in TLR9-deficient (TLR9KO) mice and wild type (WT) mice. Femoral artery ligation significantly increased RNA expression of TLR9 (20-times) in WT mice and plasma levels of single-stranded DNA and double-stranded DNA, endogenous ligands for TLR9, in both strains of mice compared with each sham-operated group (P<0.05). Laser Doppler perfusion imaging demonstrated that TLR9KO mice significantly improved the ratio of the blood flow in the ischemic to non-ischemic limb compared with WT mice at 2 weeks after ligation (P<0.05). TLR9KO mice showed less accumulation of macrophages and less expression of inflammatory molecules (e.g., TNF-α, MCP-1 and IL-1β in ischemic muscle compared with WT mice (P<0.05, respectively). In vitro experiments using thioglycolate-stimulated peritoneal macrophages demonstrated that CpG ODN, agonistic oligonucleotide for TLR9, promoted the expression of pro-inflammatory molecules (e.g., MCP-1 and TNF-α) in WT macrophages (P<0.05, respectively) but not in TLR9 KO macrophages. Furthermore, activation of TLR9 by CpG ODN inhibited migration and proliferation of endothelial cells as determined by scratch-wound assay and MTS assay, respectively (P<0.05). Conclusion: Our results suggested that TLR9 enhances inflammation and affects migration and proliferation of endothelial cells, leading to impaired blood flow recovery in ischemic limb. TLR9 may serve as a potential therapeutic target for ischemic limb disease.

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