FOXO4-knockdown suppresses oxidative stress-induced apoptosis of early pro-angiogenic cells and augments their neovascularization capacities in ischemic limbs

PLoS One. 2014 Mar 24;9(3):e92626. doi: 10.1371/journal.pone.0092626. eCollection 2014.

Abstract

The effects of therapeutic angiogenesis by intramuscular injection of early pro-angiogenic cells (EPCs) to ischemic limbs are unsatisfactory. Oxidative stress in the ischemic limbs may accelerate apoptosis of injected EPCs, leading to less neovascularization. Forkhead transcription factor 4 (FOXO4) was reported to play a pivotal role in apoptosis signaling of EPCs in response to oxidative stress. Accordingly, we assessed whether FOXO4-knockdown EPCs (FOXO4KD-EPCs) could suppress the oxidative stress-induced apoptosis and augment the neovascularization capacity in ischemic limbs. We transfected small interfering RNA targeted against FOXO4 of human EPCs to generate FOXO4KD-EPCs and confirmed a successful knockdown. FOXO4KD-EPCs gained resistance to apoptosis in response to hydrogen peroxide in vitro. Oxidative stress stained by dihydroethidium was stronger for the immunodeficient rat ischemic limb tissue than for the rat non-ischemic one. Although the number of apoptotic EPCs injected into the rat ischemic limb was greater than that of apoptotic EPCs injected into the rat non-ischemic limb, FOXO4KD-EPCs injected into the rat ischemic limb brought less apoptosis and more neovascularization than EPCs. Taken together, the use of FOXO4KD-EPCs with resistance to oxidative stress-induced apoptosis may be a new strategy to augment the effects of therapeutic angiogenesis by intramuscular injection of EPCs.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adult
  • Aged
  • Animals
  • Apoptosis*
  • Cell Cycle Proteins
  • Cytokines / metabolism
  • Female
  • Forkhead Transcription Factors / deficiency*
  • Forkhead Transcription Factors / genetics
  • Gene Expression Regulation / genetics
  • Gene Knockdown Techniques*
  • Hindlimb / blood supply*
  • Humans
  • Ischemia / genetics
  • Ischemia / pathology
  • Ischemia / physiopathology*
  • Male
  • Neovascularization, Physiologic
  • Oxidative Stress*
  • Phenotype
  • Rats
  • Reactive Oxygen Species / metabolism
  • Transcription Factors / deficiency*
  • Transcription Factors / genetics

Substances

  • Cell Cycle Proteins
  • Cytokines
  • FOXO4 protein, human
  • FOXO4 protein, rat
  • Forkhead Transcription Factors
  • Reactive Oxygen Species
  • Transcription Factors

Grants and funding

This work was supported by the Grant-in-Aid from the Strategic Research Foundation for Private Universities from MEXT, Japan (http://www.mext.go.jp), to Kurume University; the Grant-in-Aid for Young Scientists (B) [23791509] to T.N; and the Grant from Kimura Memorial Heart Foundation (http://kimura-kinen.com/contents/zaidan/index.html) to T.N. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.