Injectable multifunctional microgel encapsulating outgrowth endothelial cells and growth factors for enhanced neovascularization

Abstract

Recent cell-based therapy approaches have employed both nanotechnologies and other biomedical technologies to enhance their therapeutic potential. A combined strategy using therapeutic stem/progenitor cells and angiogenic proteins is attractive for the treatment of vascular disease. In this study, we developed an injectable multifunctional micro-sized gel system (microgel), composed of arginine-glycine-aspartic acid (RGD)-conjugated alginate, for the delivery of both cells and growth factors in vivo. The microgels encapsulated with outgrowth endothelial cells (OECs) and growth factors (vascular endothelial growth factor, VEGF, and hepatocyte growth factor, HGF) were formed via electrospraying. Cells encapsulated within the microgel exhibited a time-dependent proliferation with enhanced cell viability, and the size-controlled microgels resulted in sustained release of growth factors for enhanced new vessel formation by tube formation and rat aorta sprouting in vitro. Increased angiogenesis was also estimated in mice treated with RGD-microgel containing OECs and growth factors. Furthermore, injection of the multifunctional microgel into a hindlimb ischemia model improved blood flow perfusion and increased the capillary density by histological analysis. Compared with hydrogel system, injectable microgel system was shown to be superior with no toxicity. Overall, our injectable multifunctional microgel system can be attributed to deliver potential therapeutic agents/cells for the treatment of vascular diseases.

Biomek Cell Workstation: A Flexible System for Automated 3D Cell Cultivation

“…These include human chondrocytes, 22 dental-derived mesenchymal stem cells (MSCs), 23 embryonal stem cell (hESC) lines (Endeavor-1 and -2; MEL-1), 24 human hepatocellular carcinoma cell lines (MHCC97L, HCCLM3), 25 human hepatoma cell line HepG2, 26 and endothelial cells (OECs). 12 Spheroid cultures are primarily formed from cancer cell lines: human hepatoma cell line HepG2 (ATCC HB-8065), MCF-7 (ATCC HTB-22), 15 cell lung cancer (NSCLC) cell lines (A549, Colo699), 16 MDA-MB-231 breast cancer cells, HeyA8 ovarian cancer cells, and human mammary green fluorescent protein (GFP)-fibroblast cell line. 17 Furthermore, we also manufactured 3D constructs in the form of spheroid cultures with cervix carcinoma cells to produce in vitro tumors without carrier material.…”

“…13 Instead, the bead formations needed different matrices to generate 3D constructs. These are alginate 12,[22][23][24][25][26] and alginate hydrogel. 23 But biocompatible sodium alginate was used in the present study.…”

“…21 The semiautomated bead formation with endothelial cells (OECs) and growth factors (vascular endothelial growth factor [VEGF], hepatocyte growth factor [HGF]) with the electrospraying procedure was investigated by Kim et al The device realized the formation using a high-voltage source associated with a syringe pump and stainless-steel nozzle, as well as a syringe needle (28 gauge) from NanoNC (NNC-ESP 200, Seoul, South Korea). 12 The manually produced spheroid cultures were realized by different labware: 60 wells of a MicroWell MiniTray (Nunc) 15 and Gravity-PLUS microtissue culture system (InSphero AG). 16 However, the automated formation of HeLa cell-spheroid cultures in this paper was realized by cell seeding using the Biomek NX liquid handler in 384-well hanging drop plates.…”

“…• Petri dish 12,[23][24][25][26] • 384-well hanging drop plates 17,18 • 60 wells of a MicroWell MiniTray (Nunc) 15 • Gravity-PLUS microtissue culture system (InSphero AG) 16 • 15 mL tubes…”

“…• Semiautomated formation by microfluidic device with a two-channel fluid jacket microencapsulator, 23 air-driven droplet, generator (Nisco Engineering), 24 electrostatic droplet generator (in-house development), 25 automated formation by Inotech IER-20 encapsulator (Inotech), 26 electrospraying method 12 • Manual 15,16 • Automated cell seeding by CyBi-Disk workstation 16,18 • Manual…”