Adipose Stromal Cells Amplify Angiogenic Signaling via the VEGF/mTOR/Akt Pathway in a Murine Hindlimb Ischemia Model: A 3D Multimodality Imaging Study

PLoS One. 2012;7(9):e45621. doi: 10.1371/journal.pone.0045621. Epub 2012 Sep 20.

Abstract

Although adipose-derived stromal cell (ADSC) transplantation has been demonstrated as a promising therapeutic strategy for peripheral arterial disease (PAD), the mechanism of action behind the observed therapeutic efficacy of ADSCs remains unclear. This study was designed to investigate the long-term outcome and therapeutic behavior of engrafted ADSCs in a murine hindlimb ischemia model using multimodality molecular imaging approaches. ADSCs (1.0×10(7)) were isolated from Tg(Fluc-egfp) mice which constitutively express dual-reporter firefly luciferase and enhanced green fluorescent protein (Fluc(+)-eGFP(+), mADSCs(Fluc+GFP+)), then intramuscularly injected into the hindlimb of BALB/c-nu mice after unilateral femoral artery ligation and excision. Abbreviated survival (∼5 weeks) of post-transplant mADSCs within the ischemic hindlimb was longitudinally monitored using noninvasive bioluminescence imaging (BLI), fluorescence imaging (FRI), and bioluminescence tomography with micro-computed tomography (BLT/micro-CT). Use of the BLT/micro-CT system enabled quantitative 3-dimensional (3D) imaging of the cells' distribution and kinetics in vivo. Engrafted mADSCs improved blood perfusion recovery, ambulatory performance and prognosis of the ischemic hindlimb, probably by inducing angiogenesis and formation of collateral vessels, which could be visualized using laser Doppler perfusion imaging (LDPI), micro-CT angiography, vascular-cast imaging, and immunofluorescence. mADSCs augmented activation of the pro-angiogenic VEGF/mTOR/Akt pathway in vivo, even though the cells failed to incorporate into the host microvasculature as functional components. Downregulation of VEGF/mTOR/Akt signaling using small molecule inhibitors counteracted mADSC-induced angiogenesis and perfusion restoration. This study demonstrates for the first time the spatiotemporal kinetics and functional survival of transplanted mADSCs in a PAD model using in vivo 3D multimodality imaging. Our study indicates that mADSCs potentiate pro-angiogenic signal amplification via a VEGF/mTOR/Akt-dependent pathway, and thereby promote recovery from hindlimb ischemia.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adipose Tissue / pathology*
  • Animals
  • Cell Transplantation
  • Hindlimb / blood supply*
  • Imaging, Three-Dimensional
  • Ischemia / metabolism*
  • Ischemia / pathology
  • Ischemia / surgery
  • Mice
  • Mice, Inbred BALB C
  • Mice, Nude
  • Mice, Transgenic
  • Neovascularization, Pathologic*
  • Proto-Oncogene Proteins c-akt / metabolism*
  • Signal Transduction*
  • Stromal Cells / pathology*
  • TOR Serine-Threonine Kinases / metabolism*
  • Vascular Endothelial Growth Factor A / metabolism*

Substances

  • Vascular Endothelial Growth Factor A
  • TOR Serine-Threonine Kinases
  • mTOR protein, mouse
  • Proto-Oncogene Proteins c-akt

Grant support

This work was supported by National Basic Research Program of China (2012CB518101); National Natural Science Foundation of China (No. 30970845, No. 81090274, No. 81090270, and No. 81100579); Innovation Team Development Grant by China Department of Education (2010CXTD01); China’s Ministry of Science and Technology 863 Program (2012AA02A603). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.