Hepatocyte Growth Factor Is a Key Paracrine Molecule Mediating the Angiogenic and Therapeutic Effects of Adipose Stromal Cells

Abstract

The use of adipose stromal cells (ASC) for promoting repair of tissues is a promising therapy, but the mechanisms are not fully understood. We previously demonstrated tissue salvage by ASCs in mouse ischemia model, and showed ASCs secrete significant level of hepatocyte growth factor (HGF) which contributes to cell survival. To determine the specific contribution of HGF, we employed knockdown of HGF using small interfering RNA (siRNA). Human ASCs were transduced with a dual-cassette lentiviral construct expressing GFP gene and either a siRNA targeting HGF (iHGF), or a control sequence. Transduction of ASCs with iHGF (ASC-iHGF) reduced HGF levels by >80% (p<0.01). The proliferation of endothelial cells increased by 1.7-fold in growth-factor-deficient minimal medium (MM) when supplemented with a 1:1 mixture of control ASC conditioned medium (CM) (p<0.01). This growth stimulation was absent when ASC-iHGF CM was added. Angiogenic sprout formation by endothelial cells was 2.0-fold higher when cultured in ASC CM compared to MM (p<0.005). This stimulation was abolished when ASC-iHGF CM was used. The effect of iHGF on in vivo potency of ASC was assayed in a mouse hindlimb ischemia model. Relative perfusion (ischemic/non-ischemic limb) by day 15 was 84+4% in mice treated systemically with control ASCs, which was significantly greater (p<0.05) than both the saline (58+6%) and ASC-iHGF (69+5%) groups. These results indicate that secretion of HGF is necessary for the potency of ASCs both in vitro and in vivo. This supports the emerging concept that local factor secretion is a key element of cell-based angiogenesis and tissue support.