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Basic science: Experiment research
e0194 Src family kinase SFK is essential for recruitment of bonemarrow progenitor cells to the ischaemic myocardium
  1. Cheng Min,
  2. Zeng Qiutang,
  3. Losordo Douglas
  1. Wuhan Union Hospital, Wuhan, China

Abstract

Background The G protein-coupled receptor CXCR4 and its ligand stromal–cell derived factor 1 (SDF-1) play an important role in directing progenitor cells (PC) homing to ischaemic tissue. The Src family protein kinases (SFK) can be activated by, and serve as effectors of, G proteins. However, whether SFK play a role in SDF-1/CXCR4-mediated PC homing is unknown.

Methods and results To investigate whether SDF-1–CXCR4 signalling activates SFK, we isolated mouse bone marrow mononuclear cells (BM MNCs) and applied onto VCAM1-coated plates, followed by addition of CXCR4 agonist SDF-1 and/or antagonist AMD3100. SDF-1 rapidly (in 2 min) and dose-dependently increased phosphorylation (activation) of Lyn, a major SFK in the BM; AMD3100 attenuated the SDF-1–induced Lyn phosphorylation. Notably, SDF-1 treatment did not increase Lyn phosphorylation in the BM MNCs isolated from Mx1-cre+CXCR4fl/fl mice in which the CXCR4 gene had been deleted. To investigate whether SFK play a role in SDF-1/CXCR4-mediated chemotaxis, we performed Boyden chamber migration assay; SU6656, a SFK inhibitor, significantly inhibited BM-MNC migration towards SDF-1 (p<0.001, n=4). To investigate whether SFK play a role in SDF-1/CXCR4-mediated BM-MNC homing to ischaemic heart tissue, we isolated BM MNCs from CXCR4BAC:eGFP transgenic mice and injected 1×106 cells into WT and SDF-1BAC:SDF1-RFP transgenic mice (in which the expression of SDF1-RFP fusion protein is driven by the SDF-1 genomic regulatory sequence and the level of total SDF-1 protein is doubled) that had undergone surgical myocardial infarction 8 h earlier. Some recipient mice also received two i.p. injections of SU6656 (6 mg/kg) at the time of cell injection and again 4 h later. We found a significantly greater amount of eGFP+ cells (1.6-fold, p<0.01, n=5) and eGFP+c-kit+ cells (1.9-fold, p<0.01, n=5) recruited in the infarct border area of the SDF-1BAC: SDF1-RFP recipients than in WT recipients. SU6656 treatments significantly reduced the amount of eGFP+ cells and eGFP+c-kit+ cells (p<0.01, n=5) in both WT and SDF-1BAC: RFP recipients and abrogate the difference between the two groups.

Conclusions SFK play a critical role in SDF-1/CXCR4–mediated BM PC homing to the ischaemic cardiac tissue thus may provide a target for modulation of tissue repair.

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