Autologous Bone-Marrow Mesenchymal Stem Cell Implantation and Endothelial Function in a Rabbit Ischemic Limb Model

Shinsuke Mikami,Kazuaki Chayama,Ayumu Nakashima,Tatsuya Maruhashi,Masahiro Nishimura,Keigo Nakagawa,Takeshi Matsumoto,Yumiko Iwamoto,Chikara Goto,Yasuki Kihara,Masato Kajikawa,Mitsuo Ochi,Kensuke Noma,Yukio Kato,Yukihito Higashi,Koichiro Tsuji

doi:10.1371/journal.pone.0067739

Abstract

BackgroundThe purpose of this study was to determine whether autologous mesenchymal stem cells (MSCs) implantation improves endothelial dysfunction in a rabbit ischemic limb model.MethodsWe evaluated the effect of MSC implantation on limb blood flow (LBF) responses to acetylcholine (ACh), an endothelium-dependent vasodilator, and sodium nitroprusside (SNP), an endothelium-independent vasodilator, in rabbits with limb ischemia in which cultured MSCs were implanted (n = 20) or saline was injected as a control group (n = 20). LBF was measured using an electromagnetic flowmeter. A total of 106 MSCs were implanted into each ischemic limb.ResultsHistological sections of ischemic muscle showed that capillary index (capillary/muscle fiber) was greater in the MSC implantation group than in the control group. Laser Doppler blood perfusion index was significantly increased in the MSC implantation group compared with that in the control group. LBF response to ACh was greater in the MSC group than in the control group. After administration of NG-nitro-L-arginine, a nitric oxide synthase inhibitor, LBF response to ACh was similar in the MSC implantation group and control group. Vasodilatory effects of SNP in the two groups were similar.ConclusionsThese findings suggest that MSC implantation induces angiogenesis and augments endothelium-dependent vasodilation in a rabbit ischemic model through an increase in nitric oxide production.

Highlights

Atherosclerotic peripheral artery disease (PAD) is associated with increased cardiovascular morbidity and mortality [1], [2]
To determine the effects of mesenchymal stem cells (MSCs) implantation on angiogenesis and endothelial function in a rabbit hind limb ischemic model, we evaluated histological, angiographical, and flow-imagical formation of capillary and limb blood flow (LBF) responses to endothelium-dependent vasodilation induced by acetylcholine (ACh) and endothelium-independent vasodilation induced by sodium nitroprusside (SNP)
MSC Culture Adherent spindle-shaped cells were seen in the cultured dish at about 4 to 5 days after initial plating (Fig. 2A)

Summary

Introduction

Atherosclerotic peripheral artery disease (PAD) is associated with increased cardiovascular morbidity and mortality [1], [2]. In the late stage of PAD, progression of tissue hypoperfusion results in ischemic ulceration and gangrene. Various angiogenic approaches, including cell therapy and gene therapy, have been tried for revascularization of ischemic tissue in animal models of ischemia and in clinical trials. Bone-marrow mononuclear cell (BM-MNC) implantation has been shown to induce therapeutic angiogenesis in both ischemic limb models and patients with limb ischemia [3,4,5]. BM-MNC implantation requires harvesting a large amount of BM under general anesthesia, which would be a burden for patients with severe complications, such as myocardial ischemia, heart failure, cerebrovascular disease, and renal failure. The purpose of this study was to determine whether autologous mesenchymal stem cells (MSCs) implantation improves endothelial dysfunction in a rabbit ischemic limb model

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