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New England Surgical Society

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Tumor Necrosis Factor-alpha Increases Paracrine Expression and Survivability of Vascular Smooth Muscle Cells
Daniel C. Sasson, Kaiti Duan, Biraja C. Dash, Henry C. Hsia
Section of Plastic and Reconstructive Surgery, Department of Surgery, Yale School of Medicine, New Haven, Connecticut, United States

Objective(s): Induced-pluripotent-stem-cell-derived-vascular smooth muscle cells (iPSC-VSMC) have been suggested as viable therapeutic agents for wound healing and revascularization through their secretion of proangiogenic factor vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). Despite much focus, methods of increasing cell paracrine secretion and survivability have thus-far yielded inconsistent results. Our objective was to determine the effect of tumor necrosis factor-alpha (TNF-α) on iPSC-VSMC survivability, VEGF expression, and bFGF expression. Design: iPSC-VSMCs from a single cell line were divided into Groups: 1 (2-D control) and 2 (TNF-α precondition). After 24-hours, TNF-α was introduced into Group 2 at a concentration of 5 ng/mL. Following a subsequent 48-hour incubation period, cells from Group 1 were divided and added to two sets of type-I collagen scaffolds of density 5mg/ml. TNF-α was then added to one of these sets (TNF-α intra-scaffold treatment). Cells from Group 2 were added to a set of scaffolds as well. After 72 hours, all scaffolds were harvested for their media, tested for cell viability, and fixed for imaging. Intra-media VEGF and bFGF levels from Group 1, Group 2, and all scaffold sets were evaluated using ANOVA testing. Results: iPSC-VSMCs exposed to TNF-α during collagen scaffold proliferation showed an increase in cell viability compared to non-exposed cells (Group 1) and TNF-α preconditioned cells (Group 2) (p < 0.0001, Figure 1). No significant difference in VEGF level was found among all three sets of scaffolds. TNF-α preconditioned cells exhibited significantly lower levels of bFGF compared to control and intra-scaffold treated sets (p = 0.0314). Preliminary imaging measurements revealed an increased number of branch points and total network area in intra-scaffold treated sets. Conclusions: These findings suggest that iPSC-VSMCs intra-scaffold TNF-α conditioning increases cell viability, while its subsequent withdrawal has the inverse effect on viability and bFGF secretion. In vivo experiments will be subsequently used to elucidate the translational healing efficacy of TNF-α in wound models.


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