Short-Term Pretreatment With Low-Dose Hydrogen Peroxide Enhances the Efficacy of Bone Marrow Cells for Therapeutic Angiogenesis

Abstract

Conclusion: Treatment of bone marrow cells (BMCs) with low-dose hydrogen peroxide (H2O2) enhances their angiogenic potency. Summary: There are currently a number of approaches and clinical trials evaluating the possibility of inducing therapeutic angiogenesis as a treatment for critical limb ischemia. Optimal delivery methods of angiogenic stimuli and the best methods of stimulating angiogenesis are being actively investigated. One possibility for stimulating angiogenesis is the use of BMCs. Both preclinical and clinical trials suggest BMCs, including stem cells and endothelial progenitor cells, can improve blood flow under ischemic conditions. Endothelial differentiation from implanted BMCs and secretion of angiogenic cytokines, such as vascular endothelial growth factor (VEGF), contribute to therapeutic angiogenesis (Nat Med 2001;7:430-6). Because H2O2 increases VEGF expression, the authors tested the hypothesis that pretreatment of BMCs with H2O2 may increase the efficacy of neovascularization using BMCs. Pretreatment of BMCs was performed with H2O2 by incubating mouse BMCs in 5μM H2O2 for 30 minutes. Both BMCs pretreated and BMCs untreated with H2O2 were studied in vivo and in vitro. Reverse transcriptase polymerase chain reaction analysis showed expression of higher levels of Flk-1 messenger RNA in H2O2 pretreated BMCs (P < .01). Production of VEGF and endothelial differentiation of bone marrow cells after 1 and 7 days of culture was also enhanced with H2O2 (P < .05). Bone marrow cells were also intramuscularly implanted into the ischemic hind limbs of mice. After 14 days of implantation, many H2O2-pretreated BMCs were viable and demonstrated endothelial differentiation and incorporation into microvessels. Survival and incorporation of untreated BMCs was relatively poor. Blood flow and microvessel density in the ischemic hind limbs was greater in mice that received BMCs pretreated with H2O2 than in those implanted with untreated BMCs (P < .05). Comment: Many approaches are being investigated for improving the efficacy of angiogenesis associated with implantation of BMC’s. Approaches using gene modification are potentially efficacious but difficult to implement. This study suggests there may be relatively simple methods for improving the therapeutic potential of angiogenesis after BMC implantation.