Subcellular Localization of Survivin Determines Its Function in Cardiomyocytes.

Rationale: Reducing cardiomyocyte death and enhancing their proliferation after myocardial infarction is perhaps the single largest challenge for cardiac tissue regeneration. Survivin (SVV) is the smallest member of the inhibitor of apoptosis (IAP) family but plays two important roles; inhibiting caspase-9 activation in the intrinsic apoptosis pathway, and regulating microtubule dynamics and chromosome segregation during cell division. Genetic depletion of cardiac SVV leads to incomplete cardiomyocyte division and abnormal heart development. However, the function of SVV in adult hearts after myocardial infarction remains unclear. Methods: A homozygous inducible cardiomyocyte-specific SVV knockout transgenic mouse model was established through crossbreeding SVVflox/flox and αMHC-MCM transgenic mice. Adult mice received consecutive intraperitoneal injection of tamoxifen to induce genetic removal of SVV in cardiomyocytes. A SVV overexpressing model was established via local delivery of SVV in wild-type mouse hearts. Results: We found that 30.82% of cardiomyocytes in the peri-infarct region of SVV knockout mice were apoptotic, significantly higher than the 22.18% in control mice. In addition, ejection fraction was 29.00±0.40% in knockout mice compared to 38.04±0.50% in control mice 21 days after myocardial infarction. On the contrary, locally overexpressing SVV in the heart improved cardiac functions. Unexpectedly, we found that altering the subcellular localization of SVV overexpression produced different outcomes. Overexpression of SVV in the cytoplasm decreased cardiomyocyte apoptosis, whereas overexpression of SVV in the nucleus enhanced cardiac regeneration. The ejection fraction of mice overexpressing SVV was 36.58±0.91%, significantly higher than 28.18±1.70% in the GFP control group. Apoptotic cardiomyocytes were only 4.63% in mouse overexpressing cytosolic SVV, compared to 9.31% in the GFP group, and activation of caspase-3 was also reduced. Moreover, mice overexpressing NLS-SVV exhibited a better ejection fraction (36.19±1.02%,) than GFP controls (26.69±0.75%). NLS-SVV enhanced H3P-positive cardiomyocytes in the border zone to 0.28%, compared to only 0.08% in GFP group, through interacting with Aurora B. Conclusions: We demonstrate the importance of SVV subcellular localization in regulating post-MI cardiac repair and regeneration. We hope that this will open new translational approaches through targeted delivery of SVV.

A nanopatterned cell-seeded cardiac patch prevents electro-uncoupling and improves the therapeutic efficacy of cardiac repair.

The heart is an extremely sophisticated organ with nanoscale anisotropic structure, contractility and electro-conductivity; however, few studies have addressed the influence of cardiac anisotropy on cell transplantation for myocardial repair. Here, we hypothesized that a graft's anisotropy of myofiber orientation determines the mechano-electrical characteristics and the therapeutic efficacy. We developed aligned- and random-orientated nanofibrous electrospun patches (aEP and rEP, respectively) with or without seeding of cardiomyocytes (CMs) and endothelial cells (ECs) to test this hypothesis. Atomic force microscopy showed a better beating frequency and amplitude of CMs when cultured on aEP than that from cells cultured on rEP. For the in vivo test, a total of 66 rats were divided into six groups: sham, myocardial infarction (MI), MI + aEP, MI + rEP, MI + CM-EC/aEP and MI + CM-EC/rEP (n ≥ 10 for each group). Implantation of aEP or rEP provided mechanical support and thus retarded functional aggravation at 56 days after MI. Importantly, CM-EC/aEP implantation further improved therapeutic outcomes, while cardiac deterioration occurred on the CM-EC/rEP group. Similar results were shown by hemodynamic and infarct size examination. Another independent in vivo study was performed and electrocardiography and optical mapping demonstrated that there were more ectopic activities and defective electro-coupling after CM-EC/rEP implantation, which worsened cardiac functions. Together these results provide comprehensive functional characterizations and demonstrate the therapeutic efficacy of a nanopatterned anisotropic cardiac patch. Importantly, the study confirms the significance of cardiac anisotropy recapitulation in myocardial tissue engineering, which is valuable for the future development of translational nanomedicine.

Instructive nanofiber scaffolds with VEGF create a microenvironment for arteriogenesis and cardiac repair.

Angiogenic therapy is a promising approach for tissue repair and regeneration. However, recent clinical trials with protein delivery or gene therapy to promote angiogenesis have failed to provide therapeutic effects. A key factor for achieving effective revascularization is the durability of the microvasculature and the formation of new arterial vessels. Accordingly, we carried out experiments to test whether intramyocardial injection of self-assembling peptide nanofibers (NFs) combined with vascular endothelial growth factor (VEGF) could create an intramyocardial microenvironment with prolonged VEGF release to improve post-infarct neovascularization in rats. Our data showed that when injected with NF, VEGF delivery was sustained within the myocardium for up to 14 days, and the side effects of systemic edema and proteinuria were significantly reduced to the same level as that of control. NF/VEGF injection significantly improved angiogenesis, arteriogenesis, and cardiac performance 28 days after myocardial infarction. NF/VEGF injection not only allowed controlled local delivery but also transformed the injected site into a favorable microenvironment that recruited endogenous myofibroblasts and helped achieve effective revascularization. The engineered vascular niche further attracted a new population of cardiomyocyte-like cells to home to the injected sites, suggesting cardiomyocyte regeneration. Follow-up studies in pigs also revealed healing benefits consistent with observations in rats. In summary, this study demonstrates a new strategy for cardiovascular repair with potential for future clinical translation.