Background Vascular endothelial growth factor (VEGF) is an angiogenic growth factor that plays a critical role in several diseases, including cancer, rheumatoid arthritis and diseases of the eye. Persistent regulation of VEGF by expression of small interfering RNAs targeting VEGF represents a potential future strategy for treatment of such diseases. As a step toward this goal, the present study combines the potency of VEGF‐targeted miRNA mimics, produced from a miRNA cluster, with delivery by adeno‐associated virus (AAV)‐based vectors. Methods Nine different engineered tri‐cistronic miRNA clusters encoding anti‐VEGF effectors were generated and tested in adult human retinal pigment epithelial (ARPE‐19) cells using Renilla luciferase screening, quantitative reverse transcriptase‐polymerase chain reaction (RT‐PCR), western blotting and immunostaining analysis. In vivo efficacy was tested by the injection of scAAV2/8 vectors expressing the most effective miRNA cluster into murine hindlimb muscles, followed by quantitative RT‐PCR. Results Plasmids containing anti‐VEGF miRNA clusters showed efficient silencing of VEGF and demonstrated a combined gene silencing effect for miRNA clusters composed of multiple miRNA‐mimicked RNA interference effectors. The most potent molecule, miR‐5,10,7, resulted in a knockdown of VEGF by approximately 75%. Injection of scAAV2/8 vectors expressing miR‐5,10,7 into murine hindlimb muscles, resulted in a 44% reduction of endogenous VEGF. Conclusions We have developed miRNA clusters encoding anti‐VEGF effectors and shown, in a mouse model, that VEGF is efficiently down‐regulated by scAAV2/8‐delivered miRNA clusters, allowing potent attenuation of VEGF. These findings may contribute to the development of gene therapy based on AAV‐mediated delivery of miRNA clusters. Copyright © 2012 John Wiley & Sons, Ltd.

Autologous fat grafting and implant surgery are used for volume restoration in plastic surgery. With the aim of producing a treatment superior to current solutions, we report a randomized, controlled, data assessor-blinded clinical trial comparing fat grafts enriched with ex vivo-expanded autologous adipose-derived stromal cells (ASCs) to nonenriched fat grafts in breast augmentation. The intervention group received ASCenriched fat grafts (≥20 × 10 6 viable ex vivo-expanded ASCs per milliliter fat), and the control group received conventional nonenriched fat grafts. Volume retention was measured by magnetic resonance imaging, and clinical photographs were taken simultaneously for outcome evaluation. ASC-enriched fat grafts had significantly higher retention rates (mean = 80.2%) compared with conventional fat grafts (mean = 45.1%). Clinical photos showed statistically significant superior results in the intervention group, assessed by independent clinical experts. These results improve the prospects for using culture-expanded ASCs in both reconstructive and cosmetic volume restoration and make the procedure an attractive alternative to conventional fat grafting and implants.

PLs from outdated platelets may be an efficient and reliable source of human growth supplement allowing for large-scale ASC expansion for clinical use.

Using anti-VEGF vectors, we have demonstrated efficient silencing of endogenous mVEGF and showed that subretinal administration of scAAV2/8-hU6-sh9 has the ability to impair vessel formation in an AMD animal model. Thus, AAV-encoded shRNA can be used for the inhibition of neovascularization, leading to the development of sustained anti-VEGF therapy.