Enhanced angiogenesis in ischemic skeletal muscle after transplantation of cell sheets from baculovirus-transduced adipose-derived stromal cells expressing VEGF165

Abstract

IntroductionCell therapy using adipose-derived stromal cells (ADSC) is an intensively developing approach to promote angiogenesis and regeneration. Administration technique is crucial and among others minimal constructs - cell sheets (CS) have certain advantages. Delivery of CS allows transplantation of cells along with matrix proteins to facilitate engraftment. Cells’ therapeutic potential can be also increased by expression of proangiogenic factors by viral transduction. In this work we report on therapeutic efficacy of CS from mouse ADSC transduced to express human vascular endothelial growth factor 165 a/a isoform (VEGF165), which showed potency to restore perfusion and protect tissue in a model of limb ischemia.MethodsMouse ADSC (mADSC) isolated from C57 male mice were expanded for CS formation (106cells per CS). Constructs were transduced to express human VEGF165 by baculoviral (BV) system. CS were transplanted subcutaneously to mice with surgically induced limb ischemia and followed by laser Doppler perfusion measurements. At endpoint animals were sacrificed and skeletal muscle was evaluated for necrosis and vessel density; CS with underlying muscle was stained for apoptosis, proliferation, monocytes and blood vessels.ResultsUsing BV system and sodium butyrate treatment we expressed human VEGF165 in mADSC (production of VEGF165 reached ≈ 25-27 ng/ml/105 cells) and optimized conditions to ensure cells’ viability after transduction. Implantation of mock-transduced CS resulted in significant improvement of limb perfusion, increased capillary density and necrosis reduction at 2 weeks post-surgery compared to untreated animals. Additional improvement of blood flow and angiogenesis was observed after transplantation of VEGF165-expressing CS indicating enhanced therapeutic potential of genetically modified constructs. Moreover, we found delivery of mADSC as CS to be superior to equivalent dose of suspended cells in terms of perfusion and angiogenesis. Histology analysis of extracted CS detected limited proliferation and approximately 10 % prevalence of apoptosis in transplanted mADSC. Significant vascularization of CS and infiltration by monocytes were found in both – BV-transduced and control CS indicating graft and host interaction after transplantation.ConclusionsDelivery of ADSC by subcutaneous transplantation of CS is effective for stimulation of angiogenesis and tissue protection in limb ischemia with a potential for efficacy improvement by BV transduction to express VEGF165.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0199-6) contains supplementary material, which is available to authorized users.

Effects of adipose stem cell sheets on colon anastomotic leakage in an experimental model: Proof of principle

“…Collagen deposition was also similar in the control and experimental groups on POD7. The lack of increased capillary density in the ASC sheet treated group conflicts with previously published data [16,17] indicating that ASCs promoted angiogenesis in animal models characterized by low blood perfusion and ischemia such as hind limb ischemia [16], myocardial infarction, and skin wounds in diabetic rats [17]. However, no such effects were seen in a dilated cardiomyopathy model [15] or in gastric ulcer healing [33].…”

“…Clearly, there is a need for new strategies therefore we propose a novel approach in which ASCs are applied to the surgical wound area in cell sheets that are permeable to growth factors secreted by the ASCs and can be handled with ease by the surgeon. Therapeutic potential of ASC sheets was shown to improve cardiac tissue regeneration in the treatment of myocardial infarction [14], dilated cardiomyopathy [15], healing of hind limb ischemia [16] and chronic non-healing skin wounds [17]. Since cell sheet technology has been successfully used in improving healing of other soft tissues (i.e., heart, skin) as we mentioned above, we explored whether ASC sheets could serve as an ideal approach for the local delivery of cells to enhance healing and prevent leakage after intestinal anastomosis.…”

Effect of Cell Seeding Density and Inflammatory Cytokines on Adipose Tissue-Derived Stem Cells: an in Vitro Study

“…This is in agreement with studies directed at in vivo use of ASC-sheets. In these studies, the use of high density ASC-sheets was linked to enhanced tissue healing and angiogenesis [10, 13, 14, 21]. In addition, our data indicate that stimulating ASC-sheets with pro-inflammatory stimuli in vitro may be beneficial regarding tissue vascularization.…”

A promising therapeutic option for diabetic bladder dysfunction: Adipose tissue‐derived stem cells pretreated by defocused low‐energy shock wave

“…However, its precise mechanisms remain unknown. Revascularization and innervation are commonly considered to be the main pathophysiological changes of wound tissues triggered by shock wave (Hayashi et al, ; Jin et al, ; Makarevich et al, ; Zhao et al, ). We previously reported that MSCs were activated in vitro by DLSW via several signalling pathways (Zhao et al, ).…”