(2010). Modification of mesenchymal stem cells for cardiac regeneration. Expert Opinion on Biological Therapy: Vol. 10, No. 3, pp. 309-319. doi: 10.1517/14712590903455997

Neovessel formation is a complex process governed by the orchestrated action of multiple factors that regulate EC specification and dynamics within a growing vascular tree. These factors have been widely exploited to develop therapies for angiogenesis-related diseases such as diabetic retinopathy and tumor growth and metastasis. WNT signaling has been implicated in the regulation and development of the vascular system, but the detailed mechanism of this process remains unclear. Here, we report that Dickkopf1 (DKK1) and Dickkopf2 (DKK2), originally known as WNT antagonists, play opposite functional roles in regulating angiogenesis. DKK2 induced during EC morphogenesis promoted angiogenesis in cultured human endothelial cells and in in vivo assays using mice. Its structural homolog, DKK1, suppressed angiogenesis and was repressed upon induction of morphogenesis. Importantly, local injection of DKK2 protein significantly improved tissue repair, with enhanced neovascularization in animal models of both hind limb ischemia and myocardial infarction. We further showed that DKK2 stimulated filopodial dynamics and angiogenic sprouting of ECs via a signaling cascade involving LRP6-mediated APC/Asef2/Cdc42 activation. Thus, our findings demonstrate the distinct functions of DKK1 and DKK2 in controlling angiogenesis and suggest that DKK2 may be a viable therapeutic target in the treatment of ischemic vascular diseases.

Mesenchymal stem cells (MSCs) therapy has limitations due to the poor viability of MSCs after cell transplantation. Integrin-mediated adhesion is a prerequisite for cell survival. As a novel anti-death strategy to improve cell survival in the infarcted heart, MSCs were genetically modified to overexpress integrin-linked kinase (ILK). The survival rate of ILK-transfected MSCs (ILK-MSCs) was augmented by about 1.5-fold and the phosphorylation of ERK1/2 and Akt in ILK-MSCs were increased by about three and twofold, respectively. ILK-MSCs demonstrated an increase of twofold in the ratio of Bcl-2/Bax and inhibited caspase-3 activation, compared with hypoxic MSCs. The adhesion rate of ILK-MSCs also had a 32.2% increase on the cardiac fibroblast-derived three-dimensional matrix and ILK-MSCs showed higher retention by about fourfold compared to unmodified MSCs. Six animals per group were used for the in vivo experiments analyzed at 1 week after occlusion of the left coronary artery. ILK-MSC transplanted rats had a 12.0% ± 3.1% smaller infarct size than MSC-treated rats after ligation of left anterior descending coronary artery. Transplantation of ILK-MSCs not only led to a 16.0% ± 0.4% decrease in the fibrotic heart area, but also significantly reduced the apoptotic positive index by two-thirds when compared with ligation only. The mean microvessel count per field in the infarcted myocardium of ILK-MSCs group was increased relative to the sham group and MSCs group. In conclusion, the ILK gene transduction of MSCs further assisted cell survival and adhesion, and improved myocardial damage when compared with MSC only after transplantation. Disclosure of potential conflicts of interest is found at the end of this article.

Mesenchymal stem cell (MSC) therapy for myocardial injury has inherent limitations due to the poor viability of MSCs after cell transplantation. In this study, we directly delivered Hsp70, a protein with protective functions against stress, into MSCs, using the Hph-1 protein transduction domain ex vivo for high transfection efficiency and low cytotoxicity. Compared to control MSCs in in vitro hypoxic conditions, MSCs delivered with Hph-1-Hsp70 (Hph-1-Hsp70-MSCs) displayed higher viability and anti-apoptotic properties, including Bcl2 increase, reduction of Bax, JNK phosphorylation and caspase-3 activity. Hsp70 delivery also attenuated cellular ATP-depleting stress. Eight animals per group were used for in vivo experiments after occlusion of the left coronary artery. Transplantation of Hph-1-Hsp70-MSCs led to a decrease in the fibrotic heart area, and significantly reduced the apoptotic positive index by 19.5 ± 2%, compared to no-treatment controls. Hph-1-Hsp70-MSCs were well-integrated into the infarcted host myocardium. The mean microvessel count per field in the infarcted myocardium of the Hph-1-Hsp70-MSC-treated group (122.1 ± 13.5) increased relative to the MSC-treated group (75.9 ± 10.4). By echocardiography, transplantation of Hph-1-Hsp70-MSCs resulted in additional increases in heart function, compared to the MSCs-transplanted group. Our results may help formulate better clinical strategies for in vivo MSC cell therapy for myocardial damage.Disclosure of potential conflicts of interest is found at the end of this article.