Enhancing In Vivo Survival of Adipose-Derived Stromal Cells Through Bcl-2 Overexpression Using a Minicircle Vector

Abstract

Tissue regeneration using progenitor cell-based therapy has the potential to aid in the healing of a diverse range of pathologies, ranging from short-gut syndrome to spinal cord lesions. However, there are numerous hurdles to be overcome prior to the widespread application of these cells in the clinical setting. One of the primary barriers to effective stem cell therapy is the hostile environment that progenitor cells encounter in the clinical injury wound setting. In order to promote cellular survival, stem cell differentiation, and participation in tissue regeneration, relevant cells and delivery scaffolds must be paired with strategies to prevent cell death to ensure that these cells can survive to form de novo tissue. The Bcl-2 protein is a prosurvival member of a family of proteins that regulate the mitochondrial pathway of apoptosis. Using several strategies to overexpress the Bcl-2 protein, we demonstrated a decrease in the mediators of apoptosis in vitro and in vivo. This was shown through the use of two different clinical tissue repair models. Cells overexpressing Bcl-2 not only survived within the wound environment at a statistically significantly higher rate than control cells, but also increased tissue regeneration. Finally, we used a nonintegrating minicircle technology to achieve this in a potentially clinically applicable strategy for stem cell therapy. STEM CELLS TRANSLATIONAL MEDICINE 2013;2:690 -702

Minicircle microporation-based non-viral gene delivery improved the targeting of mesenchymal stem cells to an injury site

“…It is speculated that the difference of transfection efficiency originates from a different MSC source (hUC vs hAT and hBM); in our experiments, the hAT-and hBM-MSCs also showed different transfection efficiencies. On the other hand, Hyun et al used nucleofection technology, instead of the cationic liposome method, and obtained 25e35% transfection efficiency in hAT-MSCs [18]. Compared with these efficiencies, we achieved much higher transfection efficiency (up to 63e66%) in hAT-and hBM-MSCs using microporation, a newly developed electroporation technology.…”

“…In the case of minicircle, the cationic liposome method also seemed to be also inefficient although it showed higher transfection efficiency compared with only plasmid. Until now, minicircles have been employed for transgene expression in MSCs in two reported studies [18,19]. Daneshvar et al transfected human umbilical cord (hUC)-MSCs with minicircles, which generated 18% GFP þ MSCs [19].…”

“…Especially, genetically engineered Escherichia coli capable of inducible in vivo recombination and following the enzymatic degradation of bacterial backbone made a great improvement in minicircle production by reducing time and labor to similar levels as those of routine plasmid DNA preparation [16]. Minicircles have been employed for various purposes including transgene expression, gene therapy, DNA vaccine, and the generation of induced pluripotent stem cells [11,12,17]; however, there have been only a few studies that used minicircles for transfection of MSCs [18,19]. In these studies, minicircles were delivered to MSCs by the conventional cationic liposome method or nucleofection technology, but the resulting transfection efficiencies were low (less than 35%).…”

Surveillance of Stem Cell Fate and Function: A System for Assessing Cell Survival and Collagen ExpressionIn Situ

“…Indeed, prior studies have shown that Bcl-2 can be used to enhance stem cell survival following transplantation into calvarial defects and stented fullthickness skin wounds. 22 Osteoblasts may represent an ideal target for such an intervention.…”

Assessment of Hematotoxicity and Genotoxicity among paint Workers in Assiut Governorate: a case control study

“…Human B-cell Leukemia/lymphoma2 (Bcl-2) and Human 8-Hydroxy-deoxyguanosine (8-OHdG) levels were determined quantitatively exactly as described by Hyun et al (2013) and Sova et al (2010) respectively. The kits supplied by Glory Science Co., Ltd. cat n. TX 78840, USA.…”