Vitronectin-Based, Biomimetic Encapsulating Hydrogel Scaffolds Support Adipogenesis of Adipose Stem Cells

Abstract

Soft tissue defects are relatively common, yet currently used reconstructive treatments have varying success rates, and serious potential complications such as unpredictable volume loss and reabsorption. Human adipose-derived stem cells (ASCs), isolated from liposuction aspirate have great potential for use in soft tissue regeneration, especially when combined with a supportive scaffold. To design scaffolds that promote differentiation of these cells down an adipogenic lineage, we characterized changes in the surrounding extracellular environment during adipogenic differentiation. We found expression changes in both extracellular matrix proteins, including increases in expression of collagen-IV and vitronectin, as well as changes in the integrin expression profile, with an increase in expression of integrins such as aVb5 and a1b1. These integrins are known to specifically interact with vitronectin and collagen-IV, respectively, through binding to an Arg-Gly-Asp (RGD) sequence. When three different short RGD-containing peptides were incorporated into three-dimensional (3D) hydrogel cultures, it was found that an RGD-containing peptide derived from vitronectin provided strong initial attachment, maintained the desired morphology, and created optimal conditions for in vitro 3D adipogenic differentiation of ASCs. These results describe a simple, nontoxic encapsulating scaffold, capable of supporting the survival and desired differentiation of ASCs for the treatment of soft tissue defects.

Adipose Stem Cells Incorporated in Fibrin Clot Modulate Expression of Growth Factors

“…Human ASCs isolated from lipoaspirate, transduced with a constitutive mCherry-luciferase reporter plasmid, and selected for by previously established methods 26 were expanded in culture, and isotyping was performed with flow cytometry to confirm the presence of stem cell markers. 26 On thaw, passage 4 to 6 cells were suspended in media consisting of 60% Dulbecco's modified Eagle's medium (DMEM, Life Technologies, Carlsbad, CA) with 10% fetal bovine serum (Atlas Biologicals, Fort Collins, CO) and 40% MesenPRO medium (Life Technologies). Cells and media were centrifuged 1,200 g for 3 minutes, and resulting cell pellets were resuspended in 3:2 DMEM þ 10% fetal bovine serum:MesenPro media and switched to 100% MesenPRO 24 to 48 hours later.…”

Strategies for Bioengineered Scaffolds that Support Adipose Stem Cells in Regenerative Therapies

“…There are Strategies for bioengineered scaffolds that support adipose stem cells in regenerative therapies Review many avenues currently under investigation to determine the best method of combining ASCs and synthetic scaffolds to obtain an optimal graft or implant for the desired application. [88]. The use of vitronectin-derived attachment peptides promoted the development of larger lipid vacuoles, further suggesting that the interaction of the ASCs with their scaffolding may have a significance impact on the desired differentiation and health of implanted cells [90,91].…”

“…Peptides considered to be highly adhesive lead to smaller lipid vacuoles and thus immature adipocytes. However, peptides containing RGD derived from vitronectin (less adhesive) allowed ASCs to attach when incorporated into the hydrogel, while remaining rounded morphologically [88]. This approach demonstrates that the initial environment encountered by ASCs may influence their ability to differentiate in 3D scaffolds.…”

Mechanophysical Cues in Extracellular Matrix Regulation of Cell Behavior

“…The ECM has been studied as a scaffold for cell culture for over 40 years . Laminin, fibronectin, collagen, and vitronectin have been adopted as cell culturing scaffolds, substrates or components. These macromolecules facilitate cell culturing and improve cell survival rates .…”