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Endothelial cells (ECs) that are differentiated from induced pluripotent stem cells (iPSCs) can be used in establishing disease models for personalized drug discovery or developing patient-specific vascularized tissues or organoids. However, a number of technical challenges are often associated with iPSC-ECs in culture, including instability of the endothelial phenotype and limited cell proliferative capacity over time. Early senescence is believed to be the primary mechanism underlying these limitations. Sirtuin1 (SIRT1) is an NAD(+)-dependent deacetylase involved in the regulation of cell senescence, redox state, and inflammatory status. We hypothesize that overexpression of the SIRT1 gene in iPSC-ECs will maintain EC phenotype, function, and proliferative capacity by overcoming early cell senescence. SIRT1 gene was packaged into a lentiviral vector (LV-SIRT1) and transduced into iPSC-ECs at passage 4. Beginning with passage 5, iPSC-ECs exhibited a fibroblast-like morphology, whereas iPSC-ECs overexpressing SIRT1 maintained EC cobblestone morphology. SIRT1 overexpressing iPSC-ECs also exhibited a higher percentage of canonical markers of endothelia (LV-SIRT1 61.8% CD31(+) vs. LV-empty 31.7% CD31(+), P

The study of biomaterials for gene delivery in tissue engineering and regenerative medicine is a growing area, necessitating the investigation of new biomaterials and gene delivery vectors. Poly(1,8-octanediol citrate) (POC) and poly(glycerol-sebacate) (PGS) are biodegradable, biocompatible elastomers that have tunable mechanical properties, surface characteristics, and degradation rate. The objective of this work was to investigate whether POC and PGS would support the immobilization and release of lentivirus to allow sustained and localized transgene expression. Porous biomaterials were prepared using salt as a porogen, and in vitro and in vivo transgene expression from immobilized and released lentiviruses were assessed. Cells seeded onto biomaterials loaded with lentiviruses yielded titer-dependent transgene expression in vitro. Lentivirus activity on both biomaterials was maintained for at least 5 days. When implanted subcutaneously in rats, POC and PGS with immobilized lentivirus exhibited sustained and localized transgene expression for at least 5 weeks. This research demonstrates that lentivirus immobilization on POC and PGS is feasible and potentially useful for a variety of tissue engineering and regenerative medicine applications. © 2012 Wiley Periodicals, Inc. J Biomed Mater Res Part A, 2013.

The synthesis of poly(diol-co-citrate) elastomers that are biocompatible with vascular cells and can modulate the kinetics of the NO release based on the diol of selection is reported. NO-mediated cytostatic or cytotoxic effects can be controlled depending on the NO dose and the exposure time. When implanted in vivo in a rat carotid artery injury model, these materials demonstrate a significant reduction of neointimal hyperplasia. This is the first report of a NO-releasing polymer fabricated in the form of an elastomeric perivascular wrap for the treatment of neointimal hyperplasia. These elastomers also show promise for other cardiovascular pathologies where NO-based therapies could be beneficial.

Since the discovery of nitric oxide (NO) in the 1980s, this cellular messenger has been shown to participate in diverse biological processes such as cardiovascular homeostasis, immune response, wound healing, bone metabolism, and neurotransmission. Its beneficial effects have prompted increased research in the past two decades, with a focus on the development of materials that can locally release NO. However, significant limitations arise when applying these materials to biomedical applications. This Feature Article focuses on the development of NO-releasing and NO-generating polymeric materials (2006-2011) with emphasis on recent in vivo applications. Results are compared and discussed in terms of NO dose, release kinetics, and biological effects, in order to provide a foundation to design and evaluate new NO therapies. Nitric oxide-releasing and -generating polymeric materials are currently under investigation for biomedical applications. Clinical complications such as thrombus formation (first panel), platelet adhesion (second panel), neointimal hyperplasia (third panel), and wound healing (last panel) may benefit from advances in this field. Recent progress in the topic is reviewed herein. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim., M.C.J. and M.C.S. contributed equally to this work. M.C.J. acknowledges support from the NIH Biotechnology Training Grant. M. C. S. is greatly indebted to the Ministerio de Ciencia e Innovacion of Spain for a postdoctoral fellowship >Juan de la Cierva>. R. L. acknowledges support from the AHA Predoctoral Fellowship. G. A. A. acknowledges support by the American Heart Association Established Investigator Award.

Bypass graft procedures are commonly performed to treat narrowing or blocked vessels. However, damage to the vascular wall from the surgical intervention can lead to pro-inflammatory events and oxidative stress that promote intimal hyperplasia and may eventually lead to restenosis. Therefore, strategies that seek to reduce inflammation and oxidative stress during the initial stages of wound healing at the graft anastomoses are expected to inhibit intimal hyperplasia and reduce graft failure rates. Sirtuin-1 (Sirt1), an NAD+-dependent lysine deacetylase, protects against inflammation and oxidative stress by deacetylating the RelA/p65 subunit of NF-kB and FOXO transcription factors. We hypothesize that the localized overexpression of Sirt1 at the perivascular wall via lentivirus entrapped in a thermoresponsive gel referred to as nanonets will lead to an improved vascular healing response at the graft anastomoses and attenuate intimal hyperplasia via reduced tissue inflammation and oxidative stress. Human aortic adventitial fibroblasts (AoAFs) were transduced with lentiviruses encoding for Sirt1 and assessed for NF-kB activity, cytokine expression, senescence, and upregulation of antioxidant genes. To assess perivascular transgene delivery, nanonets containing lentiviruses encoding GFP were applied perivascularly around the aortas of rats. After one week of in vivo transduction, aortas were explanted and probed for GFP positive cells. Upon TNFα stimulation, NF-kB activity and cytokine expression levels were decreased when compared to non-transduced and empty vector-transduced AoAFs. Furthermore, Sirt1-overexpression prevented cellular senescence and protected against oxidative stress. Nanonets containing lentiviruses effectively transduced adventitial fibroblasts in vivo. In conclusion, Sirt1 overexpression in AoAFs may be protective against inflammation and oxidative stress.